Abstract The hydrolyses of a series of p-nitrophenyl carboxylates with different acyl chains were investigated at 25 °C, pH 7.8 using cyclic dipeptides[cyclo(–l- or d-Xyz–His–) consisting of l-histidine (l-His) and an α-amino acid (Xyz) with different aliphatic side chains] as catalysts. Consequently, cyclo(–d-Leu–l-His–) and cyclo(–d-Val–l-His–) were found to be specifically effective catalysts for the hydrolysis of p-nitrophenyl laurate. These trans (d-l-type) cyclic dipeptides were much more efficient catalysts than their diastereomers, and were more reactive than imidazole, despite the fact that the former are less basic than the latter. The conformation of the cyclic peptides in solution was investigated using proton magnetic resonance spectroscopy and the relationship between conformation and catalytic activity was investigated. Consequently, it was found that the hydrophobic interaction between a catalyst and a substrate and the stereochemical fit for the cooperation of functional groups in the intramolecular nucleophilic catalysis are very important in order to attain a highly efficient catalysis. It was also found that the functional groups should have a certain size and flexibility in order to realize an effective stereochemical fit between functional groups of catalysts and substrates.