title: Oleanolic acid from antifilarial triterpene saponins of Dipterocarpus zeylanicus induces oxidative stress and apoptosis in filarial parasite Setaria digitata in vitro.
abstract: Absence of a drug that kills adult filarial parasites remains the major challenge in eliminating human lymphatic filariasis (LF); the second leading cause of long-term and permanent disability. Thus, the discovery of novel antifilarial natural products with potent adulticidal activity is an urgent need. In the present study, methanol extracts of leaves, bark and winged seeds of Dipterocarpus zeylanicus (Dipterocarpaceae) were investigated for macro and microfilaricidal activity. Two antifilarial triterpene saponins were isolated from winged seed extracts by bioactivity guided chromatographic separation and identified using Nuclear Magnetic Resonance and mass spectroscopic analysis as oleanolic acid 3-O-β-D- glucopyranoside (1) (IC50 = 20.54 μM for adult worms, 19.71 μM for microfilariae ) and oleanolic acid 3-O-α-L-arabinopyranoside (2) (IC50 = 29.02 μM for adult worms, 25.99 μM for microfilariae). Acid hydrolysis of both compounds yielded oleanolic acid (3) which was non or least toxic to human peripheral blood mono nuclear cells (Selectivity index = >10) while retaining similar macrofilaricidal (IC50 = 38.4 μM) and microfilaricidal (IC50 = 35.6 μM) activities. In adult female worms treated with 50 and 100 μM doses of oleanolic acid, condensation of nuclear DNA, apoptotic body formation and tissue damage was observed by using Hoechst 33342 staining, TUNEL assay and Hematoxylin and Eosin staining respectively. A dose dependent increase in caspase 3/CED3 activity and decrease in total protein content were also observed in these parasites. A dose dependant DNA fragmentation was observed in adult parasites and microfilariae. Decreased levels of reduced glutathione (GSH) and elevated levels of glutathione S transferase (GST), superoxide dismutase (SOD) and reactive oxygen species (ROS) were also observed in parasites treated with oleanolic acid indicating an oxidative stress mediated apoptotic event. Compound 3/oleanolic acid was thus identified as a potent and safe antifilarial compound in vitro.
authors.affiliation: Institute of Biochemistry, Molecular Biology and Biotechnology, University of Colombo, Colombo 03, Sri Lanka.
authors.forename: K S
authors.initials: KS
authors.lastname: Senathilake
authors.affiliation: Institute of Biochemistry, Molecular Biology and Biotechnology, University of Colombo, Colombo 03, Sri Lanka. Electronic address: erick@ibmbb.cmb.ac.lk.
authors.forename: E H
authors.initials: EH
authors.lastname: Karunanayake
authors.affiliation: Institute of Biochemistry, Molecular Biology and Biotechnology, University of Colombo, Colombo 03, Sri Lanka.
authors.forename: S R
authors.initials: SR
authors.lastname: Samarakoon
authors.affiliation: Institute of Biochemistry, Molecular Biology and Biotechnology, University of Colombo, Colombo 03, Sri Lanka.
authors.forename: K H
authors.initials: KH
authors.lastname: Tennekoon
authors.affiliation: Department of Chemistry, Faculty of Science, University of Colombo, Colombo 03, Sri Lanka.
authors.forename: E D
authors.initials: ED
authors.lastname: de Silva
authors.affiliation: HEJ Research Institute of Chemistry, International Center for Chemical and Biological Sciences, University of Karachi, Karachi, Pakistan. Electronic address: adikariamine@gmail.com.
authors.forename: A
authors.initials: A
authors.lastname: Adhikari
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000378:metabolism; D053938:DNA Fragmentation / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D019669:Embryophyta / Q000737:chemistry*; D005260:Female; D020287:In Situ Nick-End Labeling; D020128:Inhibitory Concentration 50; D007928:Lethal Dose 50; D009828:Oleanolic Acid / Q000302:isolation & purification / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects*; D024301:Plant Bark / Q000737:chemistry; D010936:Plant Extracts / Q000302:isolation & purification / Q000494:pharmacology; D018515:Plant Leaves / Q000737:chemistry; D017382:Reactive Oxygen Species / Q000378:metabolism; D012503:Saponins / Q000737:chemistry*; D012639:Seeds / Q000737:chemistry; D017182:Setaria Nematode / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D012719:Setariasis / Q000188:drug therapy / Q000469:parasitology; D013194:Staining and Labeling; D014315:Triterpenes / Q000737:chemistry
chemical_list: D010936:Plant Extracts; D017382:Reactive Oxygen Species; D012503:Saponins; D014315:Triterpenes; D009828:Oleanolic Acid; D053148:Caspase 3
keywords: Chromatography; Dipterocarpaceae; Elephantiasis; Filarial; Helminthes; Microfilaria; Oleanolic acid; Parasites; Setaria nematode; Triterpene