Intestinal ischemia/reperfusion (II/R) is a clinical event associated with high morbidity and mortality. AMP-activated protein kinase (AMPK), a central cellular energy sensor, is associated with oxidative stress and inflammation. However, whether the AMPK is involved in the II/R-induced intestinal injury and the underlying mechanism is yet to be elucidated. Propofol has a protective effect on organs; yet, its specific mechanism of action remains unclear. This study explored the role of the AMPK-Sirt1-autophagy pathway in intestinal injury, and whether propofol could reduce intestinal injury and investigated the mechanisms in a rat model of II/R injury as well as a cell model (IEC-6 cells) of hypoxia/reoxygenation (H/R). Propofol, AMPK agonist (AICAR) and AMPK inhibitor (Compound C) were then administered, respectively. The histopathological changes, cell viability and apoptosis were detected. Furthermore, the levels of proinflammatory factors, the activities of oxidative stress, diamine oxidase, and signaling pathway were also analyzed. The results demonstrated that the AMPK-Sirt1-autophagy pathway of intestine was activated after II/R or H/R. Propofol could further activate the pathway, which reduced intestinal injury, inhibited apoptosis, reversed inflammation and oxidative stress, and improved the 24-hour survival rate in II/R rats in vivo, and attenuated H/R-induced IEC-6 cell injury, oxidative stress, and apoptosis in vitro, as fine as changes in AICAR treatment. Compound C abrogated the protective effect of propofol on II/R and H/R-induced injury. These results suggested a crucial effect of AMPK on the mechanism of intestinal injury and might provide a new insight into the mechanism of propofol reducing II/R injury.