The influence of drug thermodynamic activity and niosome composition, size, lamellarity and charge on the (trans)dermal delivery of tretinoin (TRA) was studied. For this purpose, tretinoin was incorporated at saturated and unsaturated concentrations in both multilamellar (MLV) and unilamellar (UV) vesicular formulations using two different commercial mixtures of alkyl polyglucosides: octyl-decyl polyglucoside and decyl polyglucoside. Positively and negatively charged vesicular formulations were prepared using either stearylamine or dicetylphosphate as a charge inducer. Niosomes made with polyoxyethylene (4) lauryl ether and liposomes made with soy phosphatidylcholine were also prepared and studied. Vesicular formulations were characterised by transmission electron microscopy and optical and light polarized microscopy for vesicle formation and morphology, and by dynamic laser light scattering for size distribution. The effect of the vesicular incorporation of tretinoin on its (trans)dermal delivery through the newborn pig skin was also investigated in vitro using Franz cells, in comparison with a commercial formulation of the drug (RetinA). The amount of tretinoin delivered through and accumulated in the several skin layers was detected by HPLC. Overall, obtained results showed that tretinoin cutaneous delivery is strongly affected by vesicle composition and thermodynamic activity of the drug. In particular, small, negatively charged niosomal formulations, which are saturated with tretinoin, have shown to give higher cutaneous drug retention than both liposomes and commercial formulation. Moreover, interactions between skin and vesicles seem to depend on physico-chemical properties of the main component of the vesicular bilayer.