Exercise and hypoxia can independently generate free radicals subsequent to a decrease in intracellular oxygenation (Bailey et al., 2007). The oxidative stress typically associated with exercise-induced arterial hypoxaemia (Bailey et al., 2001) may have important implications for vascular homeostasis since the superoxide anion can directly scavenge endothelium-derived nitric oxide (NO) (Gryglewsky et al., 1986). Thus, the present study tested the hypothesis that exercise-induced arterial hypoxaemia would be associated with a “downstream” reduction in the systemic bioavailability of NO. Eighteen healthy male volunteers aged 26 ± 6 (mean ± SD) years were examined at rest in normoxia, after 6h passive exposure to normobaric hypoxia that required volunteers to inspire 12% oxygen (HYP-REST) and immediately following a maximal cycling test to volitional exhaustion (HYP-EX). The venous plasma (200μL injection) and red blood cell (RBC-100μL injection) concentrations of total NO were assayed using a modified ozone-based chemiluminescence technique (Rogers et al., 2005) and not subject to plasma volume correction. Arterial haemoglobin oxygen saturation (SaO2) was measured using pulse-oximetry calibrated at the respective inspirates. Data were not normally distributed (Shapiro-Wilk-W tests) and thus differences relative to the normoxic control values were assessed using Wilcoxon Matched Pairs Signed Ranks Tests. Despite a marked decrease in SaO2 during HYP-REST (-12 ± 4%, P< 0.05 vs. normoxia), no changes were observed in the plasma (-6.4 ± 201.4 nmol/L, P > 0.05) or RBC (+15.6 ± 121.6 nmol/L, P> 0.05) concentration of total NO. In contrast, HYP-EX was associated with a more marked reduction in SaO2 (-18 ± 6%, P < 0.05) and concomitant decrease in plasma (-155.5 ± 228.1 nmol/L, P< 0.05) and RBC (-45.0 ± 119.2 nmol/L, P < 0.05) total NO. The present findings indicate that active but not passive exposure to inspiratory hypoxia caused a reduction in the systemic bioavailability of NO. We suggest that this may be due to the more severe arterial hypoxaemia encountered when exercise is superimposed on inspiratory hypoxia and its subsequent ability to compound reductive stress (Bailey et al., 2001).