The cellular uptake and toxicity of a number of substituted 3-nitronaphthalimides was investigated. Uptake of these compounds into cells was initially rapid and reached a plateau after several hours, where in some cases intracellular concentrations were much greater than the corresponding extracellular concentrations. Little uptake was obtained, however, with a compound carrying an acidic substituent. Toxicity studies divided the compounds into two main groups; those where survival curves were convex and those where survival curves were concave. The shapes of survival curves of the latter group did not appear to reflect depletion of extracellular drug. Uptake and toxicity of different drugs were not well correlated and bioreductive metabolism of the nitro-substituent did not appear to be a major contributor to toxicity. There was no consistent differential toxicity of these drugs in aerobic and hypoxic conditions. It was concluded that the nature of the ring substituent had more effect on toxicity than the absolute concentration of the naphthalimide ring or bioreductive metabolism of the nitro-group.