Myocardial ischemia-reperfusion (I/R) injury continues to be a significant clinical challenge, and ferroptosis has been identified as a major contributing factor to its development. Puerarin (Pue), an isoflavone derived from , has demonstrated promising cardioprotective effects, although the mechanisms involved are not fully elucidated. This study explores Pue's ability to attenuate myocardial I/R damage through the modulation of ferroptosis, specifically focusing on the role of the 14-3-3η protein. Network pharmacology identified 356 potential targets for Pue, with 25 genes overlapping between myocardial I/R injury and ferroptosis pathways. Molecular docking analysis revealed a strong interaction between Pue and 14-3-3η, suggesting a mechanistic link. studies using H9c2 cardiomyocytes showed that Pue pretreatment significantly improved cell viability and reduced lactate dehydrogenase (LDH) release under conditions of anoxia/reoxygenation (A/R). Pue's inhibition of ferroptosis was evidenced by reduced iron accumulation, decreased malondialdehyde (MDA) levels, lowered reactive oxygen species (ROS), and boosted anti-oxidant defenses. Central to these protective effects was the upregulation of 14-3-3η, and knockdown experiments confirmed its pivotal role in ferroptosis regulation. Furthermore, Pue preserved mitochondrial function by stabilizing mitochondrial membrane potential, limiting mitochondrial permeability transition pore (mPTP) opening, and improving mitochondrial energy metabolism and structural integrity. These activities were all mediated by 14-3-3η. , Pue administration in a rat I/R model significantly reduced myocardial injury markers and improved cardiac function, and its effects were reversed when 14-3-3η expression was downregulated. This study provides compelling evidence that Pue mitigates myocardial I/R injury by inhibiting ferroptosis through 14-3-3η modulation, and presents a novel therapeutic avenue for cardioprotection.