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Our objective was to determine the optimal amino acid (aa) formula to mitigate the effect of radiation on the electrolyte and nutrient absorption and mucosal barrier functions of the gastrointestinal (GI) tract.Studies were conducted on small intestinal mucosa of irradiated NIH Swiss mice (0, 1, 3, 5 or 7 Gy). The mucosal barrier mechanism was studied for dilution potential, conductance, and plasma endotoxin and IL‐1β; electrolyte, glucose, and aa absorption were studied with Ussing chamber flux studies and transepithelial electrical measurements. Amino acids that improved GI functions were used to formulate a drink that was tested in survival studies.Radiation (0 vs 7 Gy) decreased anion selectivity (0.48±0.05 vs 0.91±0.07) and increased paracellular conductance (30.5±4.1 vs 54.0±10.1). Plasma endotoxin levels were elevated at 7 Gy (20 vs 120). Radiation (0 vs 7 Gy) decreased Na absorption (9.8±0.8 vs − 0.6±0.3) and increased electrogenic Cl secretion (10.1±1.5 vs 0.1±0.01). Glucose absorption exhibited a dose‐dependent decrease and was abolished in 7‐Gy mice (140.7±32.2 vs 3.2±0.8). Lysine, aspartic acid, glycine, isoleucine, threonine, tyrosine, valine, serine, and tryptophan improved GI functions. Survival studies using the aa mixture showed increased survival for lethally irradiated mice.The aa mixture improved GI mucosal function and mitigated radiotoxicity.Support provided by Enterade, LLC.