Although hyperbaric oxygen (HBO) therapy has been reported to help heal chronic foot ulcers in patients with diabetes mellitus (DM), production of HBO-related oxidative stress is a concern. To assess the therapeutic effect and oxidative stress of HBO, a 2-week, prospective, randomized, controlled clinical study was conducted from January 1, 2010 to January1, 2012 among 36 consecutively admitted patients with diabetic foot ulcers (DFU). Average patient age was 60.08 ± 5.97 years and average DM duration was 16.4 ± 11.3 years; 86.1% had type 2 DM, and 47.2% had Wagner grade-III foot ulcers. Patients randomized to the control group (n = 18) received standard care including offloading, wound debridement, and glucose control. HBO treatment group patients (n = 18) received standard care and twice-daily HBO sessions for 90 minutes at 2.5 atmospheres absolute (ATA) 5 days a week for 2 weeks. Transcutaneous oxygen pressure (TcPo2) at the edge of the ulcer and wound size were measured at baseline and after 7 and 14 days of treatment. Ulcer tissues were harvested on days 7 and 14 to determine oxidative stress by measuring malondialdehyde (MDA) and antioxidant enzyme (superoxide dismutase [SOD], catalase [CAT], and glutathione peroxidase [GPx]) levels. Compared to baseline, TcPo2 in the HBO group increased on day 7 (477.8 ± 118.2 mm Hg versus 37.06 ± 5.23 mm Hg, P <0.01) and day 14 (501.1 ± 137.7 mm Hg versus 35.61 ± 4.85 mm Hg, P <0.01). Ulcer size reduction in the HBO group was greater than that of the control group (42.4% ± 20.0% versus 18.1% ± 6.5%, P <0.05). MDA levels, SOD, and CAT were all significantly higher in the HBO than in the control group on day 14 (P<0.05). The results of this study suggest HBO treatment for 2 weeks initiates a healing response in chronic DFUs, but the observed oxidative stress in local ulcer tissue may offset this effect long-term. Until needed additional research has been conducted, prolonged and/or inappropriate HBO treatment should be avoided.