Development of resistance by the malaria parasite Plasmodium falciparum has created challenges in the eradication of this deadly infectious disease. Hence newer strategies are adopted to combat this disease and simultaneously new lead/hit identification is going on worldwide to develop new chemotherapeutic agents against malaria.In this study, 44 flavonoids found mainly in the fruit juice of Citrus species having traditional use in malaria-associated fever were selected for in silico multiple-target directed screening against three vital targets of the parasite namely dihydrooroate dehydrogenase (PfDHODH), dihydrofolate reductase thymidine synthase (PfDHFR-TS) and plasma membrane P-type cation translocating ATPase (PfATP4) to find out new lead molecule(s).The in silico screening was carried out using different protocols of the Biovia Discovery Studio 2018 software and Network analyzer plugin of Cytoscape 3.6.0 followed by in vitro screening of the best lead.After the screening, CF8 or luteolin was found to have good binding affinity against PfDHODH and PfATP4 with -CDocker energy 42.2719 and 33.1447 with respect to their co-crystal ligands. These findings were also supported by Structure-based Pharmacophore, DFT (Density Functional Theory) study and finally by in vitro screening of the lead with IC50 value 8.23 µm and 12.41 µm against 3D7 (chloroquine-sensitive) and RKL-9 (chloroquine-resistant) strain of P. falciparum respectively.Our study has found out a moderately active lead molecule with the predicted mode of action which can be utilized to design some new derivatives with more safety and efficacy by targeting the two enzymes.
