The effect of temperature on skeletal muscle ATP turnover, pulmonary oxygen uptake and single fibre ATP and PCr content was studied during intense cycling exercise. Six healthy male subjects performed 6-min intense (Δ50%LT-VO(2peak)) cycling, at 60 rpm, under conditions of normal (N) and elevated muscle temperature (ET). Muscle biopsies obtained from the vastus lateralis at rest, 2 and 6 min were analysed for homogenate ATP, PCr, lactate and glycogen, allowing estimation of anaerobic ATP turnover. Freeze-dried single fibres from biopsies were characterised according to their myosin heavy chain composition (type I, IIA or IIAX) and analysed for ATP and PCr content. Pulmonary gas exchange was measured throughout. There was no difference in pulmonary oxygen uptake between the trials. The elevation of muscle temperature resulted in a lower (P < 0.05) PCr content, higher (P < 0.05) lactate content and greater (P < 0.05) anaerobic ATP turnover after 2 min of exercise. There was no effect of temperature on these measures at 6 min. In single fibres it was observed that in ET, there was a lower (P < 0.05) PCr content in type I fibres after 2 min with no differences between conditions after 6 min. The present study demonstrates that elevation of muscle temperature results in a greater anaerobic ATP turnover and type I fibre PCr degradation during the initial 2 min of intense exercise.