Procambarus clarkii and Procambarus zonangulus are two of the most widespread crayfish species in North America. In regions where their ranges overlap species composition can vary greatly. The physiological basis for this variable species composition is unknown. Temperature and oxygen level are two parameters that vary in shallow water habitats. We examined the metabolic rate and hemocyanin binding affinities in relation to thermal history. Temperature acclimation did not have the predicted effect on metabolic rate. Acclimation to high temperature (30 °C) decreased metabolic rate at 35 °C for both species. Low temperature acclimation (10 °C) resulted in 20% mortality in P. clarkii and 100% mortality in P. zonangulus when exposed to 35 °C. The range of P. clarkii is known to extend farther south than that of P. zonangulus, and this response may be a consequence of adaptations to higher temperatures in this range. Hemocyanin binding affinity was directly affected by assay and acclimation temperature. The highest P50 values were recorded for crayfish of both species acclimated to 10 °C and assayed at 30 °C. There was also a shift in the isoelectric points of hemolymph proteins (possibly due to structural changes) that correlated with and an increase in the hemocyanin binding affinity following acclimation to high temperatures (30 °C) in both species. © 2006 Elsevier Inc. All rights reserved.