This study assessed the relationship between differentiated ratings of perceived exertion (RPE) and heart rate with oxygen uptake (VO2) during two graded exercise tests (GXT) to exhaustion on a cycle ergometer in 49 men and women (19-50 years) of high and low fitness. The study also assessed whether sub-maximal RPE values elicited during the GXTs could provide appropriate estimates of maximal aerobic power (VO2max) Peripheral RPE (RPEP) was higher than overall RPE (RPEO) at exhaustion in both groups (P<0.001), but the reliability of the terminal RPEO was higher (0.75 and 0.40, respectively). Fitness did not moderate the relationship of RPEO and RPEP with VO2 during the GXTs (P>0.05). However, the correlation for RPEP and VO2 was higher for women compared to men (0.98 and 0.96, respectively, P<0.05), although this is of little practical significance. In both groups, RPEO was almost as highly correlated with VO2 as heart rate during GXTs terminated at exhaustion (approximately 0.955-0.980). There were no differences between predicted and measured VO2max when VO2 values were extrapolated from sub-maximal RPEO (13, 15 and 17) intensities (42.1+/-12.5, 43.4+/-11.5, 44.2+/-11.3 and 43.3+/-10.0 ml kg(-1) min(-1), respectively). However, VO2max predicted from sub-maximal RPEP intensities was significantly lower (P<0.05). In conclusion, terminal RPEO was a more reliable measure of the RPE, and provided more accurate estimates of VO2max in healthy participants of high and low fitness when elicited from sub-maximal exercise intensities.