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Increasing nitric oxide bioavailability via supplementation with nitrate-rich beetroot juice (BR) has been shown to attenuate the negative effect of hypoxia on peripheral oxygen saturation and exercise tolerance.We investigated the effects of a single dose of concentrated BR on the physiological responses to submaximal exercise and time trial (TT) performance in trained cyclists exposed to moderate simulated altitude (approximately 2500 m).Nine competitive amateur male cyclists (age, 28 ± 8 yr; V˙O2peak at altitude, 51.9 ± 5.8 mL·kg−1·min−1) completed four exercise trials consisting of an initial graded test to exhaustion and three performance trials on a cycle ergometer. The performance trials comprised 15 min of submaximal steady-state exercise at 60% maximum work rate and a 16.1-km TT. The second and third trials were preceded by ingestion of either 70 mL of BR or nitrate-depleted BR (PLA) 3 h before exercise.Plasma nitrate (PLA, 39.1 ± 3.5 µM; BR, 150.5 ± 9.3 µM) and nitrite (PLA, 289.8 ± 27.9 nM; BR, 678.1 ± 103.5 nM) measured immediately before exercise were higher after ingestion of BR compared with that after PLA (P < 0.001, P = 0.004). V˙O2 during steady-state exercise was lower in the BR trial (2542 ± 114 mL·min−1) than that in the PLA trial (2727 ± 85 mL·min−1, P = 0.049). TT performance was significantly faster after BR (1664 ± 14 s) than that after PLA (1702 ± 15 s, P = 0.021).A single dose of BR lowered V˙O2 during submaximal exercise and enhanced TT performance of trained cyclists in normobaric hypoxia. Consequently, ingestion of BR may be a practical and effective ergogenic aid for endurance exercise at altitude.