Multi-lamellar liposomes (MLLs), prepared by shearing a lamellar phase composed of lipids (phosphatidylcholine) and surfactant (Tween 80®), were designed to control their size, charge and elasticity, the key parameters known to influence liposomes penetration through skin. Their size was tuned by the water content of the sheared lamellar phase, and by the surfactant-to-lipid ratio as was their elasticity. Their charge was varied by the incorporation of DPPG and DOTAP to confer a high negative or positive zeta potential, respectively. Couples of MLLs differing solely in one physicochemical parameter, the others kept constant, were compared to discriminate the influence of the key parameters on their penetration through a synthetic membrane, Strat-M™. Using confocal Raman microscopy, the kinetics of MLLs penetration was established for 40 h using a Franz cell dispositive under non-occlusive conditions. From these comparisons, we showed that their transversal diffusion cannot be predicted by one sole parameter but depends on a combination of their physicochemical characteristics that were enlightened. Two types of liposomes designed for topic and systemic diffusion and tested on dog-excised skin exhibited the predicted behavior. Eventually, a mechanism supported by complementary TEM analysis is proposed to shed light on MLLs skin penetration.