ABSTRACT Objective This study evaluated various biological parameters in subjects exposed to hypoxia, to verify effective monitoring for early hypoxia detection. Background Hypoxic-hypoxia is a life-threatening condition, but no method of early detection in flight has been established. Method Six healthy male subjects underwent 2 experiments at different oxygen concentrations (O2) during hypoxic exposure on different days. Subjects inhaled approximately 21% O2 for 3 minutes, hypoxia using approximately 14% O2 in Experiment 1 and 7% O2 in Experiment 2 for 3.5 minutes using a reduced oxygen breathing device. During the experiments, pupil diameter (PD), cerebral oxygenated hemoglobin (O2Hb), peripheral tissue oxygen saturation (SpO2), heart rate (HR), and blood pressure (BP) were measured. Results All 6 subjects completed both Experiments 1 and 2. ΔO2Hb showed significant correlations with %HR (r = −0.6005, P < .0001), %PD (r = −0.5227, P = .0001), and %SpO2 (r = 0.8281, P < .0001) in Experiment 1. ΔO2Hb also showed significant correlations with %HR (r = −0.7904, P < .0001), %PD (r = −0.7933, P < .0001), and %SpO2 (r = 0.7240, P < .0001) in Experiment 2. Rapid hypoxic exposure caused increase in %PD and decrease in ΔO2Hb, followed by immediate increase in %HR, and then decrease in %SpO2. Conclusion The present study suggested that monitoring of HR, PD, and O2Hb were earlier parameters to detect hypoxia than SpO2.