The mechanism of antimalarial action of the ruthenium-chloroquine complex [RuCl₂(CQ)]₂ (1), previously shown by us to be active against CQ-resistant strains of P. falciparum and P. berghei, has been investigated. This compound is stable in aqueous solution, binds to hematin and inhibits aggregation to �-hematin at pH ~ 5 to a slightly lower extent than chloroquine diphosphate (CQDP); more importantly, the heme aggregation inhibition activity of complex 1 is significantly higher than that of CQ when measured at the interface of n-octanol/aqueous acetate buffer mixtures under acidic conditions modeling the food vacuole of the parasite. Partition coefficient measurements confirm that complex 1 is considerably more lipophilic than CQ in n-octanol/water mixtures at pH ~ 5. This suggests that the principal target of complex 1 is the heme aggregation process, which has been recently reported to be fast and spontaneous at or near water/lipid interfaces. Furthermore, the activity of complex 1 against CQ-resistant strains of P. falciparum is probably related to its greater lipophilicity, in line with previous reports indicating a lowered ability of some membrane proteins to promote the efflux of highly lipophilic drugs. The metal complex also interacts with DNA by intercalation, to a comparable extent and in a similar manner to uncomplexed CQ and therefore DNA binding does not appear to be an important part of the mechanism of antimalarial action in this case.