Quinoline antimalarials decrease the rate of β-hematin formation

The strength of inhibition of β-hematin (synthetic hemozoin or malaria pigment) formation by the quinoline antimalarial drugs chloroquine, amodiaquine, quinidine and quinine has been investigated as a function of incubation time. In the assay used, β-hematin formation was brought about using 4.5 M acetate, pH 4.5 at 60 °C. Unreacted hematin was detected by formation of a spectroscopically distinct low spin pyridine complex. Although, these drugs inhibit β-hematin formation when relatively short incubation times are used, it was found that β-hematin eventually forms with longer incubation periods (<8 h for chloroquine and >8 h for quinine). This conclusion was supported by both infrared and X-ray powder diffraction observations. It was further found that the IC50 for inhibition of β-hematin formation increases markedly with increasing incubation times in the case of the 4-aminoquinolines chloroquine and amodiaquine. By contrast, in the presence of the quinoline methanols quinine and quinidine the IC50 values increase much more slowly. This results in a partial reversal of the order of inhibition strengths at longer incubation times. Scanning electron microscopy indicates that β-hematin crystals formed in the presence of chloroquine are more uniform in both size and shape than those formed in the absence of the drug, with the external morphology of these crystallites being markedly altered. The findings suggest that these drugs act by decreasing the rate of hemozoin formation, rather than irreversibly blocking its formation. This model can also explain the observation of a sigmoidal dependence of β-hematin inhibition on drug concentration.