The purpose of this investigation was to study the effect of time of day on the relationship between lactate (LT) and ventilatory thresholds (VT) of pulmonary oxygen uptake (VO₂). Seven moderately active male volunteers (26.3 ± 3.0 years, 1.74 ± 0.08 m, 76 ± 5 kg) performed a maximal incremental test (increases of 30 W every 2 min) on a cycle ergometer on consecutive days at 0900 h, 1400h and 1900 h in a randomized fashion. The anaerobic threshold was determined using both ventilatory gas analysis and blood lactate measures. Each of the following variables was recorded both at VT and the LT; heart rate (HR, beats.min^-1), minute ventilation (VE, L.min^-1), respiratory exchange ratio (RER), time to threshold (Time, sec), oxygen uptake (VO₂, ml·kg^-1.min^-1) and VO₂ as a percentage of maximal oxygen uptake (%VO₂max). The correlations between VT and LT variables analyzed by Pearson product moment correlations for each time of day. ANOVA was used to compare the data obtained at different times of the day. There were no significant differences for the data related to time of day either for ventilatory gas analysis or lactate measurements. The correlation coefficients between VT and LT variables were moderate to high (r=0.56-0.94) for time of day. However, the correlations for HR, VO₂, and %VO2max (r=0.81-0.94) were slightly stronger compared with Time, VE and RER (r=0.56-0.88). It was concluded that, the data at VT and LT were not influenced by time of day.