The purpose of this study was to investigate whether some alterations in the barrier function of normal human skin (NHS) could be reproduced in a marketed human reconstituted epidermis (REp). Acetone/ether (AE) and ethanol (ETOH) were applied onto NHS and REp as disrupting pretreatment. The influence of this application was evaluated through the assessment of the cutaneous permeability for one amphoteric drug, caffeine (CAF), transepidermal water loss (TEWL), skin impedance (SKIM), and quantification of typical epidermal lipids, the sphingolipids (SPH). Under the experimental conditions used, we observed that the barrier function of control REp was lowered as compared to that of control NHS. These discrepencies were highlighted by the pharmacokinetic results obtained with CAF. The differences observed between the two models in terms of TEWL were consistent but far less important (three- to five-fold). Surprisingly, REp lipid profile was similar to that of NHS epidermal extracts. Skin impedance results only emphasized a slight difference between the two models. Because the barrier function of REp appeared sensitive to the disrupting effect of organic solvents, we concluded that this type of skin culture might be used to predict some alterations of the human skin barrier function.