Glutamine plays a key role in the metabolism of rapidly dividing cells, including enterocytes and lymphocytes, which may contribute to its beneficial clinical effects. Gut mucosal homeostasis is achieved through a balance between cell proliferation and apoptosis. In T cells, glutamine up-regulates anti-apoptotic proteins and down-regulates pro-apoptotic proteins. In gut mucosa, glutamine prevents apoptosis in rat epithelial cell lines, whereas glutamine starvation induces apoptosis through caspase activation. Finally, glutamine specifically prevents TNF-a-related apoptosis in the human intestinal cell line HT-29. Comparative functional proteomics enable to characterize each differently expressed protein in intestinal cells in response to modifications of nutritional environment. The influence of glutamine on intestinal proteome expression in apoptotic conditions has not been studied and evaluated. This comparative proteomics study was performed in the human epithelial intestinal cell line HCT-8 under experimental apoptotic conditions to investigate the influence of glutamine on protein expression during apoptosis. The pharmaconutritional effects of glutamine were determined under 2 mM (physiological concentration) and 10 mM (pharmaconutritional concentration) conditions. About 1,800 protein spots were revealed in both conditions. Comparative assessments indicated that 28 proteins were differentially expressed significantly (i.e. at least 2-fold modulated and Student’s t-test with p=0.05) in response to an increase of glutamine concentration in the culture medium. Twenty four proteins were identified by mass spectrometry and associated databases. From these proteins, 34% are involved in cell cycle and apoptosis mechanisms, 17% in signal transduction and 13% in cytoskeleton organization. These data were integrated in a proposed schema of intestinal interactome under apoptotic conditions. In conclusion, this study provides the first holistic picture of proteome modulation by glutamine in a human enterocytic cell line under apoptotic conditions and supports further evaluation of nutritional modulation of human intestinal proteome in various pathological conditions where apoptosis may be involved.