Purpose: To evaluate the penetration of commercially available gatifloxacin and moxifloxacin into the anterior chamber of a rabbit eye using collagen shields presoaked in the antibiotics as a drug delivery system. Methods: Collagen shields, presoaked for 10 min in commercially available solutions of gatifloxacin (diluted intravenous Tequin®) or moxifloxacin (VigamoxTM) with the same concentration, were placed on the surface of each of the corneas of 12 rabbits for a total of 24 eyes (12 in each group). Aqueous humor samples were taken 3 and 6 h later. Results:The concentrations of both antibiotics were high after 3 h (3.1 ± 1.3 µg/ml for moxifloxacin and 2.3 ± 0.8 µg/ml for gatifloxacin, p = 0.22). The concentration of gatifloxacin after 6 h was statistically significantly higher than for moxifloxacin (0.76 ± 0.33 µg/ml vs. 0.29 + 0.14 µg/ml, respectively, p = 0.03). Conclusion: Our results suggest that collagen shields can be effective as a drug delivery system for the fourth-generation fluoroquinolones with a longer effect for gatifloxacin.