During procurement, isolation, and transplantation, islets are exposed to high levels of oxidative stress triggering a variety of signaling pathways that can ultimately lead to cell death. Glutamine is an important cellular fuel and an essential precursor for the antioxidant glutathione. The aim of this study was to examine the role of intraductal glutamine administration in facilitating recovery of isolated rat islets from pancreases subjected to a clinically relevant period of warm ischemia. Islets were isolated in Sprague-Dawley (SD) rats (n = 18 per group). Pancreata in groups 1 and 2 were procured immediately while groups 3 and 4 were subjected to 30-min warm ischemia. Groups 2 and 4 were treated intraductally with 5 mM glutamine prior to pancreatectomy. Exposure to 30-min warm ischemia significantly reduced islet yield [groups 1 & 2 (nonischemia): 503 +/- 29 islets/rat vs. groups 3 & 4 (ischemia): 247 +/- 26 islets/rat; p < 0.05]. Intraductal glutamine treatment significantly improved islet yield when pancreata were subjected to 30-min warm ischemia [144 +/- 16 islets/rat without glutamine (group 3) vs. 343 +/- 36 islets/rat with glutamine (group 4), p < 0.05]. Glutamine also significantly improved islet viability (values were 50 +/- 4% in group 4 vs. 27 +/- 3% in group 3, p < 0.05). Similarly, glutathione (reduced) levels were significantly elevated in both glutamine-treated groups; however, this increase was greatest in tissues exposed to ischemia (2.76 +/- 0.04 nmol/mg protein in group 4 vs. 1.66 +/- 0.04 nmol/mg protein in group 3, p < 0.05). Intraductal glutamine administration considerably improves the islet yield, viability, and augments endogenous glutathione levels in pancreata procured after a clinically relevant period of ischemia. Intraductal administration of glutamine at the time of digestive enzyme delivery into the harvested pancreas may represent a simple yet effective tool to improve islet yields in clinical isolations.