The aim of this study was to examine and compare the influence of preconditioning, perconditioning and postconditioning with creatine phosphate (PCr) on functional recovery and production of prooxidants of isolated rat heart subjected to ex vivo I-R injury on Langendorff apparatus. Wistar albino rats (male, n=40) were divided into 4 groups: control, and groups in which PCr (0.5mmol/l, 5 minutes) was perfused before (Pre group), after (Post group) or during (Per group) ex vivo induced ischemia. PCr application was associated with the great benefits on preserving cardiac contractility (in Pre group 100.96% for +(dP/dt max), 97.61.% for -(dP/dt max), in Per group 96.72% for +(dP/dt max), 95.60.% for -(dP/dt max) and in Post group 143.84% for +(dP/dt max), 104.36% for -(dP/dt max) in relation to the stabilization). In addition, PCr application prevented the rise of pro-oxidative markers during I-R injury in all therapeutic modalities. The most intensive benefits in the current investigation were observed when PCr was applied during the period of ischemia because the lowest fluctuations in the parameters of cardiac function and oxidative stress were observed. Overall, the results of this study highlight PCr-induced cardioprotection with promising prospects for future clinical use.