The nictitating membrane, corneal epithelium, and corneal stroma were investigated as drug reservoirs. A hydrophilic drug, D,L-epinephrine HCl, or a lipophilic drug, chloramphenicol, was applied topically to rabbit eyes. Tissue levels of radioactive drug-plus-metabolites and unmetabolized epinephrine were assayed up to 24 hrs later. On a per-mg-tissue basis, concentrations of epinephrine-plus-metabolites in the stroma-endothelium were similar to or higher than those in the epithelium. The percentages of radioactivity representing unmetabolized epinephrine in the stroma-endothelium were found to be similar to or higher than those in the epithelium. On a per-mg-tissue basis, concentrations of chloramphenicol-plus-metabolites were significantly higher in the epithelium than in the stroma-endothelium during the first 130 min after drug application. While the physical properties of these drugs determined whether a higher concentration was found in the epithelium or stroma-endothelium, the ninefold greater mass of the stroma-endothelium made it the major drug reservoir on a per-whole-tissue basis. The presence or absence of a nictitating membrane had little effect on the level of either drug in the epithelium, stromal-endothelium, or aqueous humor.