Some heterocyclic cations (1-methylacridinium, 1-methyl-2-hydroxyiminomethylpyridinium and 1-methyl-3-methoxy-pyridinium) cause acceleration of hydrolysis of alkyl acetates (methyl, ethyl or n-propyl acetate) by acetylcholinesterase (acetylcholine acetylhydrolase EC 3.1.1.7) (Barnett, P. and Rosenberry, T.L. (1977) J. Biol. Chem. 252, 7200-7206). In this study, it is shown that (a) other mono- and bisquaternary ligands of pyridinium, quinolinium and benzoquinolinium series accelerate methyl-, ethyl- and n-propyl-acetate hydrolysis by acetylcholinesterase, (b) these ligands generally accelerate methyl-, and ethyl- and n-propyl-acetate, -propionate and -butyrate, 2-methoxyethyl- and furfuryl-acetate, and ethylene-glycol diacetate hydrolysis by butyrylcholinesterase (acylcholine acylhydrolase, EC 3.1.1.8). At the present time, the ability to accelerate enzymatic hydrolysis of neutral substrates appears to be restricted to some heterocyclic quaternary ammonium compounds. Acceleration which occurs at physiological ionic strength (T/2 = 0.155) involves ternary enzyme-substrate-ligand complex formation and interaction of ligands with the catalytic anionic subsite. It concerns the step leading to the enzyme-substrate complex formation and/or the acylation step of enzymes. Kinetic behaviour analogy of acetylcholinesterase and butyrylcholin-esterase in the presence of the same ligands suggests that an identical acceleration mechanism arises for both enzymes.