Chitosan biopolymers are increasingly being used in advanced biomedical applications, where aqueous interactions profoundly influence their physical properties and also their in vim biomolecular and cellular activity. Here, hydration of chitosan films is studied by dielectric spectroscopy in a conventional constraining plate configuration and compared with free standing films. Film hydration proceeds by an initial water uptake followed by a spontaneous dehydration (deswelling) even in saturated atmospheres. At water contents above a critical value, similar to 9.5 wt % a dielectric loss resonance peak (beta(wet)) arises from relaxation of evolving chitosan-water complexes, below this value insufficient interchain space for oscillation of these complexes prevents beta(wet) appearing. The beta(wet) frequency was related to water content by a power law with the frequency changing by similar to 3 orders of magnitude. Importantly the scaling exponents (slopes) differed significantly for unconstrained (free standing) and volume constrained films indicating the effect of internal stresses in constrained films. Both dielectric and conductivity behavior were influenced by internal constraining stresses affecting both oscillatory motion and charge mobility. In biomedical devices, biopolymers may be free standing, surface adhered, or enclosed structures imposing different internal stresses on polymer chains and the mobility of their segments. Dielectric spectroscopy can examine these influences on dielectric and electrical characteristics, which play a critical role in biomolecular interactions. (C) 2010 Wiley Periodicals, Inc. J Appl Polym Sci 120: 1307-1315, 2011