Bioassay-Guided Isolation and Characterisation of Antiplasmodial Compounds from Cryptolepis sanguinolenta Root Collected in Cross River State, South South Nigeria

The increasing resistance of Plasmodium falciparum to frontline artemisinin-based combination therapies necessitates an urgent search for novel antiplasmodial agents from natural product sources. Cryptolepis sanguinolenta is an established ethnomedicinal plant used for malaria treatment across southern Nigeria, with several bioactive indoloquinoline alkaloids previously characterised from West African specimens. However, population-level phytochemical variation and the full spectrum of antiplasmodial constituents in Nigerian ecotypes have not been systematically evaluated. This study employs bioassay-guided fractionation to isolate and characterise the antiplasmodial compounds from root extracts of Cryptolepis sanguinolenta collected in Yakurr, Cross River State. Crude methanol extract was sequentially partitioned into hexane, ethyl acetate, and aqueous fractions. Each fraction was evaluated for in vitro antiplasmodial activity against chloroquine-sensitive (3D7) and resistant (Dd2) strains of P. falciparum using the SYBR Green I assay. The ethyl acetate fraction demonstrated the highest activity and was subjected to column chromatography and preparative HPLC. Isolated compounds were characterised by nuclear magnetic resonance (NMR), infrared spectroscopy (IR), and mass spectrometry (MS). Three known indoloquinoline alkaloids and one novel minor alkaloid were isolated and fully characterised. The novel compound exhibited an IC50 of 28 nM against Dd2, suggesting potent multi-drug resistant antiplasmodial activity. These findings support further investigation of Cross River Cryptolepis sanguinolenta as a source of novel antimalarial leads. Keywords: Cryptolepis sanguinolenta, antiplasmodial, bioassay-guided fractionation, indoloquinoline, Cross River State