Compared to murine and human hemoglobin, bovine hemoglobin has a less exothermic oxygen binding and delivers oxygen even at low temperatures. This property could improve oxygen availability for myocytes during hypothermic arrest of hearts. The aim of this study was to evaluate the advantage of using cardioplegic solutions enriched with bovine hemoglobin when storing rat hearts. Hearts excised from rats after perfusion with different cardioplegic solutions (Celsior, Celsior plus 4% human hemoglobin, Celsior plus 4% and 8% bovine hemoglobin) were compared. Biopsies were obtained from the beating hearts before cardioplegic infusion and during a 48 h period of cold storage. Adenosine triphosphate, its catabolites and markers of oxidative stress were measured as indices of preservation. The results show that bovine hemoglobin-enriched solutions highly improve adenosine triphosphate content, decreasing its catabolites; no significant changes in antioxidant status were evident. The statistically significant difference was evident up to 6 h of storage. Doubling the concentration of bovine hemoglobin produces only slight improvement. Alternative hemoglobins with different properties may improve and prolong heart storage. As bovine hemoglobin delivers oxygen even at low temperatures, it improves energy content and anabolic reactions, without decreasing oxidative stress.