The influence of stereochemistry and alkyl side chain length on the bioactivity of the Rho kinase inhibitor Y-27632 [(+)-1, R= Me] was examined by the synthesis of (+)-and (−)-1, and two alkyl chain analogs (+)-and (−)-2 (R= n-propyl) and (−)-3 (R= n-octyl) as well as their evaluation in enzymatic and neurite outgrowth assays.