Polyrotaxanes are well known as a supramolecular assembly in which many cyclic compounds are threaded onto a linear polymeric chain capped with bulky end-groups. In this paper, a polyrotaxane consisting of alpha-CDs and PEG capped with biodegradable peptide moieties was synthesized, and the interaction with stratum corneum of hairless rat skin was examined by means of a differential scanning calorimetry. The hydroxypropylated polyrotaxane was found to interact with lipid components in the stratum corneum: bound water content was significantly decreased although ordered lipid bilayers were maintained. Thus, it is suggested that our designed polyrotaxane can be feasible as novel candidates for transdermal penetration enhancers.