Hydrophobic helical peptides having nucleotide base analogues were synthesized, and the helix multilayer was formed by interlayer hydrogen bonds to investigate the surface potential of the multilayer. Hydrophobic helical peptides having a diamino-triazine group at the C-terminal were incubated with the thymine-terminated self-assembled monolayer (SAM). The thin layers of helical peptides were formed with a nearly vertical orientation. The amount of peptide adsorbed on the surface increased with increasing concentration of the peptide solution at preparation, indicating multilayer formation. The numbers of helix layers were 10 and 5 for Boc-(Leu-Aib)(8)-T (T; 4-N-(aminoethyl)amino-2,6-diamino-1,3,5-triazine) and U-Ala-(Leu-Aib)(8)-T (U; 6-methyluracil), respectively, when 0.4 mM of peptide solution was used for the preparation. The surface potentials of these multilayers were, respectively, 558 mV and 500 mV. The U-Ala-(Leu-Aib)8-T multilayer generated nearly the same surface potential as the Boc-(Leu-Aib)(8)-T multilayer, even though the membrane thickness was different. The large positive surface potential should promote electron injection from gold to the thin peptide layer, resulting in saturation of the positive potential generation.