Endoplasmic reticulum (ER) stress‑induced apoptosis serves a crucial role in the development of myocardial ischemia/reperfusion (I/R) injury. Salidroside is a phenylpropanoid glycoside isolated from Rhodiola rosea L., which is a plant often used in traditional Chinese medicine. It possesses multiple pharmacological actions and protects against myocardial I/R injury in vitro and in vivo. However, it is not yet clear whether ER stress or ER stress‑induced apoptosis contributes to the cardioprotective effects of salidroside against myocardial I/R injury. Hence, hypoxia/reoxygenation (H/R)‑treated H9c2 cardiomyocytes were used in the current study to mimic myocardium I/R injury in vivo. It was hypothesized that salidroside alleviates ER stress and ER stress‑induced apoptosis, thereby reducing H/R injury in H9c2 cells. The results demonstrated that salidroside attenuated H/R‑induced H9c2 cardiomyocyte injury, as cell viability was increased, lactate dehydrogenase release was decreased, morphological changes in apoptotic cells were ameliorated and the apoptosis ratio was reduced compared with the H/R group. ER stress was reversed, indicated by the downregulation of glucose regulated protein 78 and C/EBP homologous protein following pretreatment with salidroside. In addition, salidroside attenuated ER stress‑induced apoptosis, as the expression of cleaved caspase‑12 and pro‑apoptotic protein Bcl‑2 associated X protein and activity of caspase‑3 was decreased, while the expression of anti‑apoptotic protein Bcl‑2 was increased following pretreatment with salidroside. Furthermore, the results indicated that salidroside decreases the activation of the ER stress‑associated signaling pathway, as the expression of phosphorylated protein kinase RNA (PKR)‑like ER kinase (p‑PERK) and phosphorylated inositol‑requiring enzyme‑1α (p‑IRE1α) proteins were decreased following pretreatment with salidroside. These results demonstrate that salidroside protects against H/R injury via regulation of the PERK and IRE1α pathways, resulting in alleviation of ER stress or ER stress‑induced apoptosis in H9c2 cardiomyocytes.