Halofantrine (Hf) is a highly lipophilic antimalarial which significantly associates with triglyceride (TG) rich plasma lipoproteins, and this is likely manifest as a decrease in the free fraction of drug. This study assessed the effect of using growth media containing 10% serum containing different concentrations of TG (i.e. TG-rich plasma lipoproteins) on the IC50 of Hf determined using continuous in vitro culture of Plasmodium falciparum. Serum was collected from a human subject in either a fasted state or at various times after ingestion of a fatty meal. There was a linear and statistically significant 2.5-fold increase in the IC50 of Hf across a 6-fold range of increasing TG concentrations, with the increased IC50 values being ascribed to a decreased free fraction of Hf in the growth media due to sequestration by TG-rich lipoproteins. Chloroquine diphosphate, which is hydrophilic and not significantly bound by TG-rich lipoproteins, was used as a control and its IC50 values were independent of TG concentrations. These data indicate that consideration should be given to the adoption of standard conditions for the collection of serum with respect to pre-or postprandial states, and that subject- and disease-related factors which alter plasma lipoprotein profiles should be considered when interpreting the IC50 profile of Hf (and possibly other lipophilic antimalarials). Furthermore, although food is known to affect the pharmacokinetics of Hf, these data suggest that altered plasma lipoprotein profiles could also influence its pharmacodynamic profile.