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This study was performed to determine the effect of duration of mitomycin exposure during trabeculectomy surgery on filtration success in eyes that had and had not been subjected to previous surgery.One hundred six eyes of 92 patients were retrospectively reviewed. The effect of previous surgery and exposure duration of mitomycin 0.5 mg/ml on outcomes of trabeculectomy surgery were evaluated.The mean duration of mitomycin exposure was 0.7 ± 0.02 min (mean ± SEM) in the eyes that had not been subjected to previous surgery and 1.5 ± 0.11 min in the eyes that had (p < 0.001). The mean follow-up times were 14 months in both groups. The mean decrease in intraocular pressure was 13.6 ± 1.25 mm Hg in the group that had not been subjected to previous surgery and 13.9 ± 1.45 mm Hg in the group that had been subjected to previous surgery. Analysis of variance techniques demonstrated no predictive value of demographics, history of previous surgery, or duration of mitomycin exposure with results of trabeculectomy surgery. Hypotony was the most frequent complication in both the no previous surgery and the previous surgery groups. The incidence of complications was numerically greater in the group that had not been subjected to previous surgery.The duration of mitomycin exposure and the history of previous surgery did not correlate with postoperative intraocular pressure, medications, or incidence of complications. The exposure duration response curve of mitomycin 0.5 mg/ml in these patients appears to be flat through the time evaluated. Lower concentrations of mitomycin and shorter exposure should be used to maintain efficacy with reduced risk of complications.