ABSTRACT: INTRODUCTION:Recently, it has been speculated that protein supplementation may further augment the adaptations to chronic endurance exercise training. We assessed the effect of protein supplementation during chronic endurance exercise training on whole-body oxidative capacity (V?O2max) and endurance exercise performance. METHODS:In this double-blind, randomized, parallel placebo-controlled trial, 60 recreationally active males (age, 27 ± 6 yr; body mass index, 23.8 ± 2.6 kg·m; V?O2max, 47 ± 6 mL·min·kg) were subjected to 12 wk of triweekly endurance exercise training. After each session and each night before sleep, participants ingested either a protein supplement (PRO; 28.7 g casein protein) or an isoenergetic carbohydrate placebo (PLA). Before and after the 12 wk of training, V?O2max and endurance exercise performance (~10-km time trial) were assessed on a cycle ergometer. Muscular endurance (total workload achieved during 30 reciprocal isokinetic contractions) was assessed by isokinetic dynamometry and body composition by dual-energy x-ray absorptiometry. Mixed-model ANOVA was applied to assess whether training adaptations differed between groups. RESULTS:Endurance exercise training induced an 11% ± 6% increase in V?O2max (time effect, P < 0.0001), with no differences between groups (PRO, 48 ± 6 to 53 ± 7 mL·min·kg; PLA, 46 ± 5 to 51 ± 6 mL·min·kg; time-treatment interaction, P = 0.50). Time to complete the time trial was reduced by 14% ± 7% (time effect, P < 0.0001), with no differences between groups (time-treatment interaction, P = 0.15). Muscular endurance increased by 6% ± 7% (time effect, P < 0.0001), with no differences between groups (time-treatment interaction, P = 0.84). Leg lean mass showed an increase after training (P < 0.0001), which tended to be greater in PRO compared with PLA (0.5 ± 0.7 vs 0.2 ± 0.6 kg, respectively; time-treatment interaction, P = 0.073). CONCLUSION:Protein supplementation after exercise and before sleep does not further augment the gains in whole-body oxidative capacity and endurance exercise performance after chronic endurance exercise training in recreationally active, healthy young males.