In this study, we investigated the effect of sefsols on the skin to clarify the mechanism of the sefsol enhancement effect. In vitro percutaneous absorption experiments were performed using Franz diffusion cells. Removal of the stratum corneum and delipidization of the skin increased the permeation of diclofenac from aqueous suspension, with the enhancement effects being similar for both treatments. Further enhancement effects of diclofenac permeation by sefsol through the stripped and delipidized skin were not observed. Attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR) was employed to investigate the biophysical changes in the stratum corneum lipids by sefsols. One of the sefsols, propylene glycol mono caprylate (S-218), induced higher and broader absorbance shifts in both asymmetric and symmetric C-H bond stretching regions. However, no significant differences were observed among the sefsols with respect to peak heights and areas for both absorbances when compared with H2O treatment. These results suggest that sefsol may change the lipid-chain fluidity of the stratum corneum without lipid extraction. The accumulated amounts of diclofenac in the skin significantly increased in the presence of sefsol. Also, the amounts of diclofenac in the skin increased with the amount of sefsol in the skin. This sefsol enhancement effect was reversed at 12 h after treatment. Thus, enhancement of the diclofenac flux by sefsols is reversible and may be due to a change in the lipid-chain fluidity of the stratum corneum and improvement in drug partitioning to the stratum corneum.