The generation of reactive oxygen species is typically associated with hyperoxia and ischemia reperfusion. Recent evidence has suggested that increased oxidative stress may occur with hypoxia. We hypothesized that oxidative stress would be increased in subjects exposed to high altitude hypoxia. We studied 28 control subjects living in Lima, Peru (sea level), at baseline and following 48 h exposure to high altitude (4300 m). To assess the effects of chronic altitude exposure, we studied 25 adult males resident in Cerro de Pasco, Peru (altitude 4300 m). We also studied 27 subjects living in Cerro de Pasco who develop excessive erythrocytosis (hematocrit > 65%) and chronic mountain sickness. Acute high altitude exposure led to increased urinary F 2-isoprostane, 8-iso PGF2α (1.31 ± 0.8 μg/g creatinine versus 2.15 ± 1.1, p = 0.001) and plasma total glutathione (1.29 ± 0.10 μmol versus 1.37 ± 0.09, p = 0.002), with a trend to increased plasma thiobarbituric acid reactive substance (TBARS) (59.7 ± 36 pmol/mg protein versus 63.8 ± 27, p = NS). High altitude residents had significantly elevated levels of urinary 8-iso PGF2α (1.3 ± 0.8 μg/g creatinine versus 4.1 ± 3.4, p = 0.007), plasma TBARS (59.7 ± 36 pmol/mg protein versus 85 ± 28, p = 0.008), and plasma total glutathione (1.29 ± 0.10 μmol versus 1.55 ± 0.19, p < 0.0001) compared to sea level. High altitude residents with excessive erythrocytosis had higher levels of oxidative stress compared to high altitude residents with normal hematological adaptation. In conclusion, oxidative stress is increased following both acute exposure to high altitude without exercise and with chronic residence at high altitude. © Mary Ann Liebert, Inc.