SUMMARY: The intracellular esterases of 25 strains of Escherichia coli, growing exponentially on a minimal medium, were analysed by the acrylamide-agarose zymogram technique. Five kinds of esterase bands were defined: three major bands (A, B and C) and two minor ones. The A and B esterase bands hydrolysed α-naphthyl, β-naphythl and indoxyl acetates; they were inhibited by di-iso-fluoropropyl phosphate (DFP). Esterase band B also hydrolysed the α- and β-naphthyl butyrates and was stable at 60°C. Esterase band C hydrolysed only β-naphthyl acetate and it resisted DFP. The A, B and C esterase bands showed variations in electrophoretic mobility which seemed to indicate an intraspecific differentiation of molecular structures of the esterase that could have arisen during microbial evolution.