| Summary: | Bioprospecting native Australian plants offers the potential discovery of latent and novel bioactive compounds. The promising cytotoxic and antibacterial activity of methanolic extracts of <i>Pittosporum angustifolium</i> and <i>Terminalia ferdinandiana</i> led to further fractionation and isolation using our laboratory’s bioassay-guided fractionation protocol. Hence, the aim of this study was to further evaluate the bioactivity of the fractions and subfractions and characterize bioactive compounds using liquid chromatography mass spectroscopy (LC-MS/MS) and gas chromatography MS (GC-MS). Compounds tentatively identified in <i>P. angustifolium</i> Fraction 1 using LC-ESI-QTOF-MS/MS were chlorogenic acid and/or neochlorogenic acid, bergapten, berberine, 8′-epitanegool and rosmarinic acid. GC-MS analysis data showed the presence of around 100 compounds, mainly comprising carboxylic acids, sugars, sugar alcohols, amino acids and monoalkylglycerols. Furthermore, the fractions obtained from <i>T. ferdinandiana</i> flesh extracts showed no cytotoxicity, except against HT29 cell lines, and only Fraction 2 exhibited some antibacterial activity. The reduced bioactivity observed in the <i>T. ferdinandiana</i> fractions could be attributed to the potential loss of synergy as compounds become separated within the fractions. As a result, the further fractionation and separation of compounds in these samples was not pursued. However, additional dose-dependent studies are warranted to validate the bioactivity of <i>T. ferdinandiana</i> flesh fractions, particularly since this is an understudied species. Moreover, LC-MS/GC-MS studies confirm the presence of bioactive compounds in <i>P. angustifolium</i> Fraction 1/subfractions, which helps to explain the significant acute anticancer activity of this plant. The screening process designed in this study has the potential to pave the way for developing scientifically validated phytochemical/bioactivity information on ethnomedicinal plants, thereby facilitating further bioprospecting efforts and supporting the discovery of novel drugs in modern medicine.
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