Two populations of Hordeum leporinum have evolved resistance to paraquat within a small area in central Tasmania, Australia. One population (THL1) was more than 80-fold resistant to paraquat when treated in winter, compared with a susceptible population (THL4) collected nearby, whereas the other population (THL2) was only 19-fold resistant. Translocation of paraquat was examined in all three populations at warm and cool temperature regimes. Herbicide was applied to a basal section of the second leaf of plants kept in the dark and translocation measured after 16 h of dark and during a subsequent light period. Paraquat absorption into the treated leaf was uniformly high in susceptible and resistant populations, with >93% of the applied herbicide absorbed within 16 h in the dark at both temperatures. Translocation of paraquat out of the treated leaf was low in the dark, with <4% of the herbicide translocated to the remainder of the plant. More herbicide was translocated out of the treated leaves in susceptible plants in the dark, compared with resistant plants at both temperature regimes and more paraquat was translocated at warmer temperatures. Extensive basipetal translocation of paraquat to the rest of the plant occurred in susceptible plants following exposure of the treated plants to light. However, basipetal translocation was much reduced in resistant plants in the light and corresponded to the degree of resistance. Resistance to paraquat in H. leporinum is the result of reduced translocation of paraquat out of the treated leaves.