A high-performance liquid chromatography method for the direct separation of enantiomers of a homologous series of 1,4-disubstituted piperazine derivatives, using a chiral cellulose tris(4-methylbenzoate) stationary phase and a hexanepropan-2-ol eluent, is described. An atypical relationship was found between enantioselectivity and the carbon number of the alkyl substituent with a maximum corresponding to four-five carbon atoms in the substituent. The optimum enantioselectivity observed suggests the existence of a limited size interaction site on the stationary phase and an input to retention due to the steric exclusion of individual solutes.