Thermally induced transition between anhydrous and hydrated forms of highly crystalline β-chitin was studied by differential thermal calorimetry (DSC) and X-ray diffraction. DSC of wet β-chitin in a sealed pan gave two well-defined endothermic peaks at 85.2 and 104.7 °C on heating and one broad exothermic peak at between 60 and 0 °C on cooling. These peaks were highly reproducible and became more distinct after repeated heating-cooling cycles. The X-ray diffraction pattern of wet β-chitin at elevated temperature showed corresponding changes in d-spacing between the sheets formed by stacking of chitin molecules. These phenomena clearly show that water is reversibly incorporated into the β-chitin crystal and that the temperature change induces transitions between anhydrous, monohydrate, and dihydrate forms. The DSC behavior in heating-cooling cycles, including reversion between the two endothermic peaks, indicated that the transition between monohydrate and dihydrate was a fast and narrow-temperature process, whereas the one between the anhydrous and the monohydrate form was a slow and wide-temperature process.