Opuntia ficus indica (OFI) has many physiological effects, but a relationship between OFI and heart-rate variability (HRV) has never been established. The aim of this study was to describe the effects of a diet supplement of OFI on HRV in athletes. The first day, heart rate (HR) was measured at rest in supine (SU) and standing (ST) positions to analyze HRV in 10 athletes, followed by a randomized assignment to an OFI (5) or placebo (5) group. The next day, the athletes repeated the HRV test. One month later the crossover protocol was applied. In OFI, the high-frequency-activity HF(SU) (1,773 +/- 2,927 vs. 5,856 +/- 8,326 ms(2), p < .05), HF(ST) (295 +/- 313 vs. 560 +/- 515 ms(2), p < .05), and low-frequency LF(SU) (1,621 +/- 1,795 vs. 6,029 +/- 9,007 ms(2), p < .01) increased. HR(SU) (66 +/- 13 vs. 57 +/- 11 beats/min, p < .01) and HR(ST) (87 +/- 11 vs. 76 +/- 9 beats/min, p < .01) decreased. A diet supplement of OFI increases HF and LF activities and decreases HR.