We have developed the Rabbit Corneal Epithelial (RCE) Model to evaluate the in vitro eye irritation potential of chemicals including pharmaceuticals, cosmetics and their raw ingredients. In the model, a stratified culture of rabbit corneal epithelial cells is grown at the air–liquid interface on an amnion acting as a parabasal membrane. The alkaline exposure was restored each day in the presence of no irritants, although with the addition of sodium lauryl sulfate (SLS), which is a major irritant, the restoration of deficit was inhibited on the RCE model in a dose-dependent manner. The results of this test were comparable with those of the Draize test, and thus, this method using the RCE model may prove to be a useful and sensitive in vitro eye irritation test. The in vitro eye irritation potential of polyoxyethylene alkyl derivatives, polyoxyethylene lauryl ether (PLE), polyoxyethylene cetyl ether (PCE), polyoxyethylene stearyl ether (PSE), polyoxyethylene oleyl ether (POE), and polyoxyethylene behenyl ether (PBE) were evaluated using the RCE model containing an alkaline exposure. POE inhibited 90.2% of the restoration of deficit at a concentration of 0.5% on the 4th day after addition. Depending on the structure, an activity relationship was defined. The polyoxyethylene alkyl derivatives had distinctly different inhibitory potencies against the restoration of deficit, according to their substitution patterns. POE inhibited the restoration of deficit greater than other polyoxyethylene alkyl derivatives on the RCE model. These results indicated that the oleyl chain of POE is an important factor for inhibiting the restoration of deficit on the RCE model.