ABSTRACT Fully synthetic endoperoxide antimalarials, namely, OZ277 (RBx11160; also known as arterolane) and OZ439 (artefenomel), have been approved for marketing or are currently in clinical development. We undertook an analysis of the kinetics of the in vitro responses of Plasmodium falciparum to the new ozonide antimalarials. For these studies we used a K13 mutant (artemisinin resistant) isolate from a region in Cambodia and a genetically matched (artemisinin sensitive) K13 revertant. We used a pulsed-exposure assay format to interrogate the time dependence of the response. Because the ozonides have physicochemical properties different from those of the artemisinins, assay optimization was required to ensure that the drugs were completely removed following the pulsed exposure. Like that of artemisinins, ozonide activity requires active hemoglobin degradation. Short pulses of the ozonides were less effective than short pulses of dihydroartemisinin; however, when early-ring-stage parasites were exposed to drugs for periods relevant to their in vivo exposure, the ozonide antimalarials were markedly more effective.