Hydrations and phase transitions of ammonium, sodium and potassium salts of phosphatidylserines (PS) were investigated by small-angle X-ray scattering (SAXS) and differential scanning calorimetry (DSC). The increase in long spacings for NH4+-PS by addition of water was slightly smaller than those for the other PS salts, implying that a small amount of water penetrated into the hydrocarbon regions. The long spacings for PS salts and water system were reduced by addition of CaCl2. A mixture of phosphatidylcholine (PC) and PS in KCl aqueous solution gave a long spacing of 59 angstrom, while it separated into two long spacings at 65 and 55 angstrom in 1 mol CaCl2 solution. This SAXS result shows that Ca2+ ion induces not only two dimensional, but three dimensional phase separation of the mixture of PC and PS. The phase transition temperature of the anhydrous NH4+-PS was lowest among those of the three PS salts, but fully hydrated NH4+-PS had the highest transition temperature among them.