Freshwater turtles found in higher latitudes can experience extreme challenges to acid-base homeostasis while overwintering, due to a combination of cold temperatures along with the potential for environmental hypoxia. Histidine containing dipeptides (HCDs; carnosine, anserine and balenine) may facilitate pH regulation in response to these challenges, through their role as pH buffers. We measured the HCDs content of three tissues (liver, cardiac muscle and skeletal muscle) from the anoxia-tolerant painted turtle (Chrysemys picta bellii) acclimated to either 3 or 20 C. HCDs were detected in all tissues, with the highest content shown in the skeletal muscle. Turtles acclimated to 3 C had more HCD in their skeletal muscle than those acclimated to 20 C (carnosine = 20.8 +/- 4.5 vs 12.5 +/- 5.9 mmol/kg DM; ES = 1.59 (95%CI: 0.16 - 3.00), P = 0.013). The higher HCD content observed in the skeletal muscle of the cold-acclimated turtles suggests a role in acid-base regulation in response to physiological challenges associated with living in the cold, with the increase possibly related to the temperature sensitivity of carnosine's dissociation constant and buffering power of the skeletal muscle during anoxic submergence.