Abstract Background Nearly all amputations in people with diabetes are preceded by a foot ulcer. It has been reported that stress is an important risk factor for slower wound healing and susceptibility to infection. The purpose of this study was to objectively monitor physiological stress response in patients with diabetic foot ulcer (DFU) during a clinical visit for wound dressing change. Methods Physiological stress was continuously monitored in 20 patients (Age: 56.7±12.2 years) with DFU for duration of approximately 45min including waiting, dressing change and post-dressing period. Stress was quantified using a custom algorithm based on standard deviation of R-R intervals, heart rate variability (HRV). To identify the change in level of stress, change in HRV was compared to baseline HRV. Medium and high-stress periods were defined when HRV was in the range of 60–85% and below 60% of baseline HRV, respectively. Results During the entire time of visit medium and high stress episodes happened for 28±12.6% and 16±18% of the time respectively. Further analyses of wound dressing time revealed that medium and high stress episodes were experienced for 47±24% and 18.3±27% of dressing time respectively. In addition, the duration of medium stress was significantly increased in average by 66% (difference=21.8%, p=0.002, 95%CI [7.5, 29.5]%) during dressing change compared to entire period of clinic visit. Conclusions This pilot study shows that patients with DFU experience moderate to high stress while visiting a wound clinic, which may negatively impact wound healing outcomes. In particular, the highest stressful condition was during wound dressing change, which may be related to painful dressing or perception of pain. Further investigation is required to explore association between quantified stress and outcomes of wound healing. In addition, future studies need to explore benefit of stress management in enhancing the outcomes of wound healing in patients with diabetes.