Carbon-13 Nuclear Magnetic Resonance Investigations of Phase Transitions and Phase Equilibria in Pure and Mixed Phospholipid Bilayers

The temperature dependence of the ¹³C NMR spectra of dipalmitoylphosphatidylethanolamine (DPPE) and three lecithins-dimyristoyl-, dipalmitoyl-, and distearoylphosphatidylcholine- which have been ¹³C-labeled at the sn-2 carbonyl has been studied. In the Lg or Lp phase, an axially symmetric powder pattern of about 100-ppm breadth is observed, and this transforms to an isotropic line at the main L_β (L_β) →L_α phase transition. In the case of DPPE, this transformation occurs precipitously and, with data from 2H spectra of 2H chain labeled DPPE, is shown to be due to a change in conformation at the sn-2 carbonyl. In contrast, the 13C=0 sn-2 spectra of lecithins exhibit a gradual transformation, beginning at temperatures below the endothermic pretransition temperature. Thus, in the intermediate Pp phase, a temperature-dependent superposition of the Lp- and La-like ¹³C spectra is observed, suggesting that the Pp phase is structurally heterogeneous and exhibits properties of both the Lp and La phases. Simulation of the sn-2 ¹³C=0 powder patterns, assuming diffusion about the lipid long axis and exchange between the two conformations, provides excellent fits of the spectra. The rate constants for the exchange can be related to the lateral diffusion constants measured by other techniques, and there is good agreement between the two. Addition of cholesterol (CHOL) to the four pure lipids also results in a superposition of the Lβ (Lβ) and La patterns, and simulation of the spectra provides a means to extract the fraction of Lβ-like lipid as a function of temperature or CHOL concentration. Similar results are observed for binary mixtures of dipalmitoylphosphatidylcholine and DPPE, indicating that sn-2 carbonyl spectra should also be a useful probe for examining phase equilibria in multicomponent lipid mixtures. Collectively, the results suggest that the conformational change at the sn-2 C=0 is a general property of phospholipid phase transitions.