Microdialysis with high performance liquid chromatography and electrochemical detection (HPLC–ECD) was used to measure ascorbic acid (AA) concentrations in rabbit vitreous before and after vitrectomy. A cellulose microdialysis probe was implanted in the vitreous humor, and after stabilization, AA measurements were made daily over a 10 day period. The effect of removing two-thirds of the vitreous by vitrectomy was examined. The effect of triamcinolone acetonide (TA) was evaluated in four groups of rabbits: Group 1, sub-tenon TA (20 mg) alone; Group 2, intravitreous TA (4 mg) alone; Group 3, sub-tenon TA (20 mg) after vitrectomy, and Group 4, intravitreous TA (4 mg) after vitrectomy. The results showed that the AA concentration after vitrectomy was significantly lower from days 2 to10 with a maximum reduction of 49.5% (P<0.005) on day 7. No significant changes in the AA level was observed in Groups 1 and 2, a mild recovery of AA concentration reduction after vitrectomy was detected in Group 3. The highest recovery of the AA concentration reduction was observed in Group 4. The attenuating effect of TA treatment on the reduction of AA in the vitreous after vitrectomy was significant. This attenuating effect of the TA may be due to prevention of the disruption of the blood-aqueous barrier by its anti-inflammatory action.