Differential scanning calorimetry and X-ray diffraction of anhydrous and hydrated A-palmitoylgalactosylsphingosine (NPGS) show evidence of complex polymorphic behavior and interconversions between stable and metastable structural forms. Anhydrous NPGS exhibits three lamellar crystal forms (A, B, and B') at temperatures below 143 °C and a liquid-crystal form between 143 and 180 °C before melting to an isotropic liquid at 180 °C. The crystal B → liquid-crystal transition is accompanied by an enthalpy change, AH, of 11.2 kcal/mol of NPGS, while a relatively small enthalpy change (AH = 0.8 kcal/mol) marks the liquid-crystal→ liquid transition. The A and B' crystal forms do not hydrate readily at room temperature. When heated, crystal form A in the presence of water undergoes an exothermic transition at 52 °C to produce a thermodynamically stable hydrated crystal E form. X-ray diffraction shows that this stable bilayer crystal form has a highly ordered hydrocarbon chain packing arrangement; melting to the bilayer liquid-crystal form occurs at 82 °C with a large enthalpy change, AH = 17.5 kcal/mol of NPGS. A complex liquid-crystal → crystal transition is observed on cooling; the cooling rate independent exotherm involves the transition of the hydrated liquid crystal to an intermediate metastable crystal form identical with anhydrous crystal form A. The subsequent cooling rate dependent step involves the conversion of the metastable crystal form A to the stable crystal form E. We suggest that hydrated crystal form E is stabilized by both a highly ordered chain packing mode and a lateral intermolecular hydrogen bonding network involving the sphingosine backbone, the galactosyl group, and interbilayer water molecules. Although disruption of both the specific hydrocarbon chain packing and H-bonding networks occurs at the high enthalpy transition to the bilayer liquidcrystal La form, these two types of interactions are not reestablished simultaneously on cooling. First, recrystallization of the hydrocarbon chains accompanies removal of water from the lipid interface, leading to “dehydrated” metastable crystal form A. This is followed by a time-dependent, temperaturedependent hydration process which allows a rearrangement of the hydrogen-bonding matrix. Alterations in the NPGSNPGS and NPGS-water interactions accompany further changes in tlje hydrocarbon chain packing and lead to the formation of the stable E form.
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