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Glass transition temperature (Tg) and mechanical and water vapor barrier properties of wheat gluten films containing glycerin, sucrose, glycerin-sucrose, and glycerin-sorbitol were studied. Glycerin and sucrose were immiscible; the wheat gluten film containing both solutes showed two separate thermal transitions (-58°C and -5°C, respectively). The low temperature (low-T) transition (-58°C) was due to a glycerin-rich region. The low-T tan δ peak height influenced the tensile strength and elongation linearly and the water vapor permeability (WVP) curvilinearly. However, the Tg failed to change with solute composition and exhibited low correlation with barrier and mechanical properties. Initial addition of glycerin increased the WVP dramatically. Sucrose decreased the WVP (only slightly), but resulted in a rigid and fragile film. The gluten-glycerin-sorbitol film at a ratio of 15:3:3 (w/w), 16.7% moisture (wb), showed a single effective Tg (-42°C) and intermediate values for tensile strength, percent elongation, and WVP, which were between those of the 15:6:0 and 15:4:2 gluten-glycerin-sucrose films.