Influence of the mineralization process of collagen (Cg) by hydroxyapatite (HA) on interactions with water molecules was investigated using infrared and Raman spectroscopy and piezogravimetry methods. Hydroxyapatite was synthesized using a simplified basic reaction with and without Ti ions, followed by phase analysis with X-ray diffraction spectroscopy (XRD). Cg–HA film samples have been prepared by a process of slow evaporation of water from the solution at 4 °C. It was found that in the presence of HA some Cg hydration sites, probably corresponding to its gap region, are closed, at least partly, for direct interactions with water molecules from the Cg hydration shell. A model illustrating early mineralization process of two Cg layers in the presence of water molecules are proposed for explaining of obtained experimental data.