Recent investigation have indicated that cyclic dodeca- and tetradecapeptides, cyclo(-Leu-Orn-Leu-Orn-D-Phe-Pro)2 (Orn-DLL-12) and cyclo(-Leu-Orn-Leu-Orn-Leu-D-Phe-Pro)2 (Orn-DLL-14), which are designed on the basis of a cyclic beta-structural antibiotic, gramicidin S (GS), inhibit the growth of Gram-positive and -negative bacteria with high potency [Ando, S., Nishikawa, H., Takiguchi, H., Lee, S. & Sugihara, G. (1993) Biochim. Biophys. Acta 1147, 42-49]. In this study we designed and synthesized two analogs, Lys-DLL-12 and Lys-DLL-14, in which four Orn residues in Orn-DLL-12 and Orn-DLL-14 were replaced by Lys residues, respectively, and investigated their interactions with model membranes in terms of CD and dye-leakage experiments, antimicrobial activity and lytic activity for human erythrocytes. Both peptides newly designed showed no antimicrobial activity and no lytic activity of erythrocytes. The present CD study showed that the presence of neutral liposomes and of acidic liposomes of natural or synthetic phospholipids results in no remarkable conformational difference between Orn-DLL-12/-14. The leakage experiment showed a clear relation between the antimicrobial activity and the leakage ability in acidic synthetic phospholipid liposomes but no correlation in acidic natural ones. The difference in hydrophobic and hydrophilic balance between Orn-DLL-12/14 and Lys-DLL-12/14 (derived from the increasing hydrophobicity due to an increase of four methylene units by the substitution of Lys for Orn) may be one of the important factors in the drastic decrease in activity.