Objective: To examine the efficacy of l-alanyl-l-glutamine ingestion with a commercially available sports drink compared to the sports drink only on time to exhaustion and physiological measures during prolonged endurance exercise. Methods: Twelve endurance-trained men (23.5 ± 3.7 years; 175.5 ± 5.4 cm; 70.7 ± 7.6 kg) performed 4 trials, each consisting of a 1-hour treadmill run at 75% VO2peakfollowed by a run to exhaustion at 90% VO2peak. One trial consisted of no hydration (NHY), another required ingestion of only a sports drink (ED), and 2 trials required ingestion of a low dose (LD; 300 mg·500 ml−1) and high dose (HD) of l-alanyl-l-glutamine (1 g·500 ml−1) added to the sports drink. During the fluid ingestion trials, 250 ml was consumed every 15 minutes. Plasma glutamine, glucose, electrolytes, and osmolality were measured prior to the run (PRE) and at 30, 45, and 60 minutes. VO2, respiratory quotient (RQ), and heart rate (HR) were measured every 15 minutes. Results: Time to exhaustion was significantly longer during the LD and HD trials compared to NHY. No differences were noted in time to exhaustion between ED and NHY. Plasma glutamine concentrations were significantly elevated at 45 minutes in LD and HD trials and remained elevated at 60 minutes during HD. Sodium concentrations increased from the beginning of exercise and remained stable for the duration of the 1-hour run. At 60 minutes, plasma sodium was significantly lower in all trials compared to NHY. Conclusions: Results indicated that ingestion of the alanine-glutamine dipeptide at either the low or high dose significantly improved time to exhaustion during high-intensity exercise compared to a no-hydration trial