Tretinoin (0.1% Retin-A cream) was topically applied to human cadaver skin in vitro using Franz diffusion chambers. The photo-isomerization of tretinoin and retinoic acid percutaneous absorption in the absence of metabolic activity were assessed with and without ambient light exposure to the skin. Using HPLC, UV, and GC/MSD, a retinoid exhibiting identical chromatographic and spectral characteristics of isotretinoin was observed in the samples from the skin exposed to light, but was virtually absent in the skin samples maintained in the dark. From a single topical application of tretinoin, isotretinoin was as abundant as tretinoin in the chamber receiver solution, dermis, epidermis, and on the skin surface at 24 h after topical application. The data suggest the possibility that isotretinoin may have an important role in the pharmacology of topically applied tretinoin.