Greenhouse and laboratory studies were conducted to determine the physiological basis for selectivity of nicosulfuron and primisulfuron in 5 plant species. Differential sensitivity of the species was quantified by determining GR50 values (herbicide rate required to reduce plant growth 50%) for each species/herbicide combination. GR50 data indicated the following levels of sensitivity: corn—tolerant to both herbicides; seedling johnsongrass—sensitive to both herbicides; barnyardgrass—sensitive to nicosulfuron and tolerant to primisulfuron; giant foxtail—sensitive to nicosulfuron and tolerant to primisulfuron; and eastern black nightshade—tolerant to nicosulfuron and sensitive to primisulfuron. Studies using 14C-radiolabeled herbicides were conducted to determine whether differential herbicide absorption, translocation, or metabolism contributed to whole plant responses. Nicosulfuron and primisulfuron selectivity in corn, johnsongrass, barnyardgrass, and giant foxtail was primarily due to differential herbicide metabolism rate. Tolerant species metabolized the herbicide more rapidly and extensively than sensitive species. Differential herbicide absorption, translocation, or metabolism did not explain differential sensitivity of eastern black nightshade to the herbicides. Further studies indicated that the tolerance of eastern black nightshade to nicosulfuron and its sensitivity to primisulfuron was directly related to lower sensitivity of the acetolactate synthase (ALS) to nicosulfuron than to primisulfuron. Eastern black nightshade translocated very little (3%) of the nicosulfuron applied. The ALS sensitivity of johnsongrass and eastern black nightshade was similar in the presence of nicosulfuron. A combination of a higher ALS level and less herbicide translocation contributes to tolerance of eastern black nightshade and to sensitivity of johnsongrass to nicosulfuron.