Poly(ADP-ribose) polymerase (PARP)-1 inhibitor has been suggested to attenuate the ischemia-reperfusion injury. We investigated the protective effect of the cardioplegia with a PARP-1 inhibitor, 4-hydoxyquinazoline (4-HQ), against myocardial ischemia-reperfusion injury. Isolated rat hearts were perfused on a Langendorff apparatus and cardioplegically arrested for 90 min by perfusion with St. Thomas' Hospital solution (ST-solution). In the Group ST (n = 8), the hearts were arrested with the ST-solution alone. The Group HQ (n = 8) were treated with the ST-solution containing 4-HQ (10 µM) for cardioplegia. During reperfusion, the Group HQ showed significantly greater functional recovery of +dp/dt(max) (p = 0.005) and lower enzymatic leakage (p < 0.01). NAD(+) levels were also preserved higher in the Group HQ (p < 0.01). Immunohistochemical study revealed lesser extents of oxidative stress and apoptosis, in the Group HQ. Thus, addition of 4-HQ in the cardioplegia may provide a new intervention for myocardial protection against ischemia-reperfusion injury by decreasing NAD(+) consumption and suppressing oxidative stress.