The stable unilamellar vesicles formation, having large potential applications in biological as well as biomedical fields, has been investigated in aqueous solution composed of a zwitterionic surfactant, N-hexadecyl-N,N-dimethylammonio-1-propanesulfonate (SB-16), and water-soluble cationic poly(amino acid), poly-L-lysine (PLL). Dynamic light scattering (DLS), transmission electron microscopy (TEM), and other optical spectroscopic techniques revealed the transformation of SB-16 micelles in aqueous solutions into stable unilamellar vesicles above a certain concentration (0.008 to 0.1% w/v) of PLL. Solvation and rotational dynamics of coumarin 480 (C-480) give the information on hydration behavior around the headgroup regions of SB-16 micelle and SB-16/PLL vesicle. It was observed that the hydration nature around the headgroup regions of SB-16/PLL vesicular system is higher than the head group regions of micellar system. Thus, PLL permits more water molecules in the headgroup regions of vesicular system.