Differential scanning calorimetry (DSC) and X-ray diffraction techniques have been used to investigate the structure and thermotropic properties of synthetic, non-hydroxy fatty acid (16:0) ceramide (NFA(C16)CER) as a function of hydration. Anhydrous NFA(C16)CER shows a single, broad endothermic transition at 95.4 degrees C (delta H = 10.4 kcal/mol). On hydration, a broad exothermic transition appears at approximately 50-70 degrees C while the main endothermic transition decreases to 90.0 degrees C (delta H = 13.8 kcal/mol). The enthalpy of the exothermic transition increases with hydration to a maximum value, delta H = 4.8 kcal/mol. This polymorphic phase behavior depends on the low temperature incubation time and prior cooling rate. X-ray diffraction of fully hydrated NFA(C16)CER at 26 degrees C, shows a well-ordered lamellar phase with a bilayer periodicity d = 46.9 A. At 68 degrees C, above the first exothermic transition, X-ray diffraction shows again a lamellar phase with reduced bilayer periodicity d = 41.8 A and an increased number of both lamellar and wide-angle reflections indicative of enhanced layer and chain packing order, respectively. At 90.0 degrees C, above the main transition, the diffraction pattern shows a broad, intense reflection at 29.9 A and a diffuse reflection at 4.6 A, indicative of a melted chain phase. On cooling, NFA(C16)CER exhibits polymorphic phase behavior involving the conversion of the melted chain phase to a metastable bilayer phase. On heating, this metastable phase undergoes an exothermic transition to a stable bilayer phase; on further heating, NFA(C16)CER converts endothermically to the melted-chain phase.(ABSTRACT TRUNCATED AT 250 WORDS)