The purpose of this study was to examine the physiological determinants of endurance cycling time trial (TT) performance in a heterogeneous group of competitive male road cyclists. About 15 male cyclists who had all competed in cycling the preceding season were tested for the anthropometric variables height, body weight, leg length, ankle circumference, and body fat percentage. They were also tested for maximal oxygen consumption (VO2max), lactate threshold (LT), metabolic cost of cycling (CC), peak power output and average power output during a 30-second Wingate test, 1 repetition maximum and peak power in half squats, and a TT test on an ergometer. Heart rate and cadence (rounds per minute, RPM) were continuously measured during all cycle tests. Pearson Bivariate correlation tests and single linear regression tests were performed to obtain correlation coefficients (r), effect size (F), standard error of estimate (SEE), and 95% confidence interval. The single variable that correlated best with TT performance was power output at LT (r = 0.86, p < 0.01). Standard error of estimate was 7.5%. Lactate threshold expressed in %VO2max did not correlate significantly with TT performance. An equation representing both aerobic and anaerobic endurance capacity TT(w) = 0.95 ([VO2max/CC] TT%VO2max) + 0.05 (Wingate average) correlated strongly with TT laboratory performance (r = 0.93, p < 0.01, SEE = 5.7%). None of the strength, power, or anthropometric variables correlated significantly with TT laboratory performance.