The effects of benzalkonium chloride (BAK) on the living rabbit cornea were studied by in vivo Tandem scanning confocal microscopy (TSCM) and confirmed by conventional scanning electron microscopy (SEM). Two drops of saline or phosphate-buffered saline (PBS) containing BAK in concentrations of 0.02, 0.01, and 0.005% was applied to rabbit eyes 15 times at 5-min intervals. The solutions were pH 5.5-5.9 (saline) and pH 7.5 (PBS), and osmolarity was 275-280 (saline) and 300-307 mOsm (PBS). Immediately after application of 0.02 and 0.01% BAK, no normal corneal superficial epithelial cells could be imaged by in vivo TSCM. No swelling of the superficial epithelial cells was observed for the control solution without BAK; however, there was a small amount of desquamation. Application of as little as 0.005% BAK caused the superficial epithelial cells to swell and desquamate. The observed desquamation of corneal superficial epithelial cells increased with higher BAK concentrations applied to the eye. One hour after final drug application, inflammatory cells appeared on the surface of the cornea treated with 0.02% BAK. These findings were correlated with SEM observations. Based on the results of this study, we believe that BAK used frequently can produce clinical corneal toxicity and that the cytotoxicity of any topical ophthalmic solutions can be evaluated by in vivo TSCM.
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