Surfactant nanocarriers have received considerable attention in the last several years as interesting alternative to classic liposomes. Different pH-sensitive vesicular colloidal carriers based on Tween 20 derivatives, obtained after functionalization of the head groups of the surfactant with natural, or simply modified, amino acids, were proposed as drug nanocarriers. Dynamic light scattering, Small Angle X-ray Scattering, Trasmission Electron Microscopy and fluorescence studies were used for the physico-chemical characterization of vesicles and mean size, size distribution, zeta potential, vesicle morphology and bilayer properties were evaluated. The pH-sensitivity and the stability of formulations, in absence and in presence of foetal bovine serum, were also evaluated. Moreover, the contact between surfactant vesicles and liposomes designed to model the cellular membrane was investigated by fluorescence studies to preliminary explore the potential interaction between vesicle and cell membranes. Experimental findings showed that physico-chemical and technological features of pH-sensitive vesicles were influenced by the composition of the carriers. Furthermore, proposed carriers are able to interact with mimetic cell membrane and it is reasonable to attribute the observed differences in interaction to the architectural/structural properties of Tween 20 derivatives. The findings reported in this investigation showed that a deep and extensive physico-chemical characterization of the carrier is a fundamental step, according to the evidence that the knowledge of nanocarrier properties is necessary to translate its potentiality to in vitro/in vivo applications.