The effect of chlorhexidine acetate on the corneal penetration of sorbitol was evaluated in vitro using the enucleated pigmented rabbit cornea mounted in a modified Ussing chamber. Sorbitol penetrated the cornea poorly when compared with arnolol, a beta blocker. Sorbitol penetration was improved 85% by 0.01% chlorhexidine acetate, 2.9 times by 0.1% EDTA, and 9.6 times by stripping the corneal epithelium prior to the start of the experiment. By comparison, 0.01% chlorhexidine acetate and stripping the corneal epithelium improved the corneal penetration of arnolol only 30% and 74%, respectively, whereas stripping the corneal epithelium did not affect the corneal penetration of chlorhexidine acetate. Collectively, the above findings indicate that changes in corneal integrity may dramatically affect the corneal penetration of some inert excipients in ophthalmic formulations. Such a possibility must be considered carefully in the selection of excipients.