A 2 Â 2 factorial design was performed to determine the effect of a permeation enhancer (oleic acid/propylene glycol), iontophoresis (2 V), and the combination of the two treatments on the permeation enhancement of a model peptide, LHRH (luteinizing hormone releasing hormone), through human epidermal membrane (HEM). In parallel studies, TEAB (tetraethylammonium bromide, a small ionic solute) and sucrose (an electroosmotic flow marker) were also investigated. Structural changes in the HEM were monitored via conductance measurements, differential scanning calorimetry (DSC), and infrared (IR) spectroscopy experiments. LHRH enhancement due to enhancer in combination with iontophoresis (I þ E; 29.5 times passive permeability, P), was greater than during iontophoresis alone (I; 14.3) and enhancer treatment alone (E; 3.5). I þ E had an additive effect of I and E, indicating the mechanisms of action of the individual enhancement strategies were likely to be located at different sites in the skin. Also, no synergistic enhancement was observed with I þ E for either TEAB or sucrose. For TEAB, permeability enhancement due to I ($1400) was much higher than that due to E (14.9), and no additive effect could be detected. For sucrose, E had no effect on either passive or iontophoretic permeability, eliminating the possibility that electroosmosis could explain increases in LHRH permeability. Evidence of synergy between E and I was found, with conductance measurements indicating that I þ E synergistically increased the membrane permeability to conducting ions (Na þ and Cl À ). It appears these pathways were not available for transport for the solutes used in the current study. DSC and IR investigations showed significant changes in stratum corneum lipid structure following E treatment but not following I. These findings probably arise from the localized action of iontophoresis compared with the bulk action of enhancer. In summary, increased LHRH delivery through HEM in vitro can be achieved using an enhancer in combination with iontophoresis.