These studies were conducted to compare the effects on systemic O₂ transport of chronically vs. acutely increased Hb O₂ affinity. O₂ transport during maximal normoxic and hypoxic [inspired PO₂ (PI_O2) = 70 and 55 Torr, respectively] exercise was studied in rats with Hb O₂ affinity that was increased chronically by sodium cyanate (group 1) or acutely by transfusion with blood obtained from cyanate-treated rats (group 2). Group 3 consisted of normal rats. Hb O₂ half-saturation pressure (P₅₀; Torr) during maximal exercise was ~26 in groups 1 and 2 and ~46 in group 3. In normoxia, maximal blood O₂ convection (O_2 max = cardiac output × arterial blood O₂ content) was similar in all groups, whereas in hypoxia O_2 max was significantly higher in groups 1 and 2 than in group 3. Tissue O₂ extraction (arteriovenous O₂ content/arterial O₂ content) was lowest in group 1, intermediate in group 2, and highest in group 3 (P < 0.05) at all exercise PI_O2 values. In normoxia, maximal O₂ utilization (O_2 max) paralleled O₂ extraction ratio and was lowest in group 1, intermediate in group 2, and highest in group 3 (P < 0.05). In hypoxia, the lower O₂ extraction ratio values of groups 1 and 2 were offset by their higher O_2 max; accordingly, their differences in O_2 max from group 3 were attenuated or reversed. Tissue O₂ transfer capacity (O_2 max/mixed venous PO₂) was lowest in group 1 and comparable in groups 2 and 3. We conclude that lowering Hb P₅₀ has opposing effects on O_2 max and O₂ extraction ratio, with the relative magnitude of these changes, which varies with PI_O2, determining O_2 max. Although the lower O₂ extraction ratio of groups 2 vs. 3 suggests a decrease in tissue PO₂ diffusion gradient secondary to the low P₅₀, the lower O₂ extraction ratio of groups 1 vs. 2 suggests additional negative effects of sodium cyanate and/or chronically low Hb P₅₀ on tissue O₂ transfer.