The relationship between antimalarial activity and drug accumulation of chloroquine and amodiaquine was evaluated with four chloroquine-resistant and two chloroquine-susceptible isolates of Plasmodium falciparum. Susceptibility of the strains to amodiaquine was correlated with susceptibility to chloroquine (r2 = 0.96). Similarly, accumulation of amodiaquine was correlated with accumulation of chloroquine (r2 = 0.94). Accumulation of both chloroquine and amodiaquine was significantly reduced in chloroquine-resistant isolates (p < 0.005). For the panel of isolates, the accumulation ratio of both drugs was inversely proportional to drug susceptibility (r2 = 0.963 and 0.994 for amodiaquine and chloroquine, respectively). Time course studies highlighted a reduced initial rate of amodiaquine accumulation in chloroquine-resistant isolates compared with chloroquine-susceptible isolates, with no evidence of an enhanced drug efflux rate. Daunomycin, a modulator of parasite chloroquine transport, significantly increased steady state accumulation of both drugs in chloroquine-resistant isolates and, to a lesser extent, in chloroquine-susceptible isolates. Furthermore, daunomycin increased the initial rate of accumulation of amodiaquine in both chloroquine-resistant and chloroquine-susceptible isolates. Resistance to 4-aminoquinoline drugs is associated with reduced drug permeability rather than enhanced cellular exit of preaccumulated drug, and daunomycin seems to increase the permeability of parasites to aminoquinolines. A new model of 4-aminoquinoline resistance is proposed to take account of these and earlier observations.