["The Journal of Physiology, EarlyView. ", "\nAbstract figure legend Co‐ingesting fructose with glucose increases exogenous carbohydrate oxidation at sea level, but its effect at altitude is unclear. This study compared glucose–fructose (GLU–FRU), glucose alone (GLU) and placebo during 120 min of running at 70% V̇O2max${{\\dot{V}}_{{{{\\mathrm{O}}}_2}{\\mathrm{max}}}}$ at 2500 m in eight males. GLU–FRU increased exogenous carbohydrate oxidation and reduced endogenous carbohydrate use compared with GLU. Both GLU and GLU–FRU improved 5 km running performance compared with placebo, with no difference between carbohydrate conditions. Created in https://BioRender.com.\n\n\n\n\n\n\n\n\n\nAbstract\nThe ingestion of fructose with glucose has been shown to enhance exogenous carbohydrate oxidation during exercise at sea level compared with glucose alone. Whether this synergistic effect occurs at altitude is yet to be established. This study compared glucose–fructose co‐ingestion with the ingestion of glucose alone on fuel use and running performance at 2500 m. Ingestion of 1.2 g min−1 glucose plus 0.8 g min−1 fructose (GLU–FRU) was compared with 1.2 g min−1 glucose (GLU) and a placebo during 120 min running at 70% V̇O2max${{\\dot{V}}_{{{{\\mathrm{O}}}_2}{\\mathrm{max}}}}$ performed by eight males at 2500 m altitude. Exogenous (individual and combined contributions from fructose and glucose) and endogenous oxidation were quantified using indirect calorimetry and 13C stable mass isotope tracing techniques. Performance was measured using a 5 km treadmill time trial. Total cholesterol (CHO) oxidation for the 120 min of running was significantly higher for GLU compared with placebo, whereas fat oxidation was no different between conditions. GLU–FRU led to significantly greater exogenous CHO oxidation and lower endogenous CHO oxidation compared with GLU. Time to complete the 5 km run was significantly faster in GLU and GLU–FRU compared with placebo, but there were no differences between GLU and GLU–FRU. In conclusion, the co‐ingestion of GLU–FRU at altitude increased exogenous and spared endogenous carbohydrate oxidation over glucose alone, as seen at sea level. However, this did not provide a performance benefit for GLU–FRU. Therefore, this is the first demonstration of the synergistic effect of the co‐ingestion of GLU–FRU on exogenous CHO oxidation during running at altitude.\n\n\n\n\n\n\n\n\n\nKey points\n\nThis study is the first to compare glucose alone with glucose–fructose co‐ingestion during prolonged running at moderate altitude (2500 m).\nCombining glucose and fructose significantly increases exogenous carbohydrate oxidation and reduces reliance on internal (endogenous) carbohydrate stores compared with glucose alone.\nDespite improved fuel use, performance in a 5 km run was not better with glucose–fructose compared with glucose alone at moderate altitude (2500 m), though both were faster than placebo.\nThese findings support glucose–fructose supplementation as an effective strategy to decrease reliance on endogenous carbohydrate stores compared with glucose alone for endurance athletes training or competing at moderate altitude.\n\n\n"]