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A porcine model was used to investigate the feasibility of using VIS-NIR spectroscopy to differentiate between degrees of ischemia-reperfusion injury in the small intestine. Ten pigs were used in this study and four segments were created in the small intestine of each pig: (1) control, (2) full arterial and venous mesenteric occlusion for 8 h, (3) arterial and venous mesenteric occlusion for 2 h followed by reperfusion for 6 h, and (4) arterial and venous mesenteric occlusion for 4 h followed by reperfusion for 4 h. Two models were built using partial least square discriminant analysis. The first model was able to differentiate between the control, ischemic, and reperfused intestinal segments with an average accuracy of 99.2% with 10-fold cross-validation, and the second model was able to discriminate between the viable versus non-viable intestinal segments with an average accuracy of 96.0% using 10-fold cross-validation. Moreover, histopathology was used to investigate the borderline between viable and non-viable intestinal segments. The VIS-NIR spectroscopy method together with a PLS-DA model showed promising results and appears to be well-suited as a potentially real-time intraoperative method for assessing intestinal ischemia-reperfusion injury, due to its easy-to-use and non-invasive nature.