The WPI-NaCas-GLY antimicrobial film takes full advantage of the controlled release of active or antimicrobial agents as well as demonstrates a great potential for functioning as an alternative biodegradable polymer in practical applications. The moisture sorption kinetics of the film as an important carrier of active agents was investigated at various relative humidities (RH). The results indicated that the moisture sorption characterization and procedure of this film can be described well by the empirical Peleg model with higher confidence and concordance. The model could predict the film's moisture content at any time (Mt), the time to reach any given level of R (tR), the equilibrium moisture at any RH condition (Me), and isotherm trend based upon experimental data and modeled constants k1, k2, a, b, c, and d without giving consideration to their physical meaning. The water vapor transmission rate of the WPI-NaCas-GLY antimicrobial film increased exponentially with increasing RH due to its hydrophilicity, which was primarily caused by the presence of glycerol in a higher content. The results also suggested that aw predominately affects the film's Me values compared with the temperature factor by fixed nonlinear multiple regression analyses.