The aim of this study was to examine the variability of the oxygen uptake (VO2) kinetic response during moderate- and high-intensity treadmill exercise within the same day (at 06:00, 12:00 and 18:00 h) and across days (on five occasions). Nine participants (age 25 +/- 8 years, mass 70.2 +/- 4.7 kg, VO2max 4137 +/- 697 ml x min(-1); mean +/- s) took part in the study. Six of the participants performed replicate 'square-wave' rest-to-exercise transitions of 6 min duration at running speeds calculated to require 80% VO2 at the ventilatory threshold (moderate-intensity exercise) and 50% of the difference between VO2 at the ventilatory threshold and VO2max (50% delta; high-intensity exercise) on 5 different days. Although the amplitudes of the VO2 response were relatively constant (coefficient of variation approximately 6%) from day to day, the time-based parameters were more variable (coefficient of variation approximately 15 to 30%). All nine participants performed replicate square-waves for each time of day. There was no diurnal effect on the time-based parameters of VO2 kinetics during either moderate- or high-intensity exercise. However, for high-intensity exercise, the amplitude of the primary component was significantly lower during the 12:00 h trial (2859 +/- 142 ml x min(-1) vs 2955 +/- 135 ml x min(-1) at 06:00 h and 2937 +/- 137 ml x min(-1) at 18:00 h; P < 0.05), but this effect was eliminated when expressed relative to body mass. The results of this study indicate that the amplitudes of the VO2 kinetic responses to moderate- and high-intensity treadmill exercise are similar within and across test days. The time-based parameters, however, are more variable from day to day and multiple transitions are, therefore, recommended to increase confidence in the data.