The study investigates the influence of surface charge, generation and concentration of poly(amidoamine) (PAMAM) dendrimers on skin permeation of a model hydrophilic drug, 5-fluorouracil (5FU). Permeation studies were performed using excised porcine skin in a Franz diffusion cell. Saturated concentration of 5FU in isopropyl myristate was applied on the skin after pretreatment with dendrimers and (14)C labeled 5FU was analyzed using a liquid scintillation counter. The influence of dendrimer surface charge (G4-NH(2), G3.5-COOH, and G4-OH), generations (G2-G6-NH(2)) and concentration (0.1-10 mM of G4-NH(2)) on skin permeation of 5FU were studied. The enhancement in drug permeability coefficient (K(p)) was in the following decreasing order G4-NH(2) > G4-OH > G3.5-COOH. Dendrimer increased the skin permeation by increasing the skin partitioning of 5FU. Transepidermal water loss, skin resistance measurements and ATR-FTIR studies revealed that cationic dendrimers act by interacting with the skin lipid bilayers. Increase in G4-NH(2) concentration did not proportionally increase the skin permeation of 5FU. The 5FU K(p) was inversely related to the molecular weight of the dendrimer. G2-NH(2) dendrimer reduced skin resistance to a greater extent than higher generation dendrimers. Overall, the study showed that lower generation cationic dendrimer is more effective in enhancing the skin permeation of hydrophilic drugs.