Biological activity of the alpha-acetoxy derivatives of methylbenzylnitrosamine was evaluated using tester strains of Salmonella typhimurium. Compound III, oxidized on the benzyl moiety, was more toxic and more mutagenic than Compound IV, oxidized on the methyl side chain. Compound IV was weakly toxic and was non-mutagenic at the concentrations tested. The presence or absence of the uvrB repair system had no effect on the toxicity of either Compound II or IV. Neither the alpha-oxidized compounds nor the parent nitrosamine reverted the frameshift mutants. As a mutagen, Compound III was approximately as active as N-nitroso compounds not requiring metabolic activation, more active than alpha-acetoxy dialkylnitrosamines and less active than the cyclic alpha-acetoxynitrosopyrrolidine.