A C-13 AND H-2 NUCLEAR-MAGNETIC-RESONANCE STUDY OF PHOSPHATIDYLCHOLINE CHOLESTEROL INTERACTIONS - CHARACTERIZATION OF LIQUID-GEL PHASES

A detailed study on the structure, dynamics, and thermodynamic behavio r of phosphatidylcholine/cholesterol (PC/CHOL) mixtures was undertaken using differential scanning calorimetry (DSC) and solid-state nuclear magnetic resonance (NMR) spectroscopy. DSC thermograms of mixtures of cholesterol (CHOL) with 1,2-dipalmitoyl-sn-phosphatidylcholine (DPPC) , 1,2-distearoyl-sn-phosphatidylcholine (DSPC), and 1,2-diarachidoyl-s n-phosphatidylcholine (DAPC) showed a broadening of the first-order ge l --> liquid crystalline transition and a decrease in the transition e nthalpy, indicating a gradual loss of cooperativity for high CHOL conc entrations. DPPC and DSPC were labeled with C-13 at the carbonyl group of the sn-2 chain and H-2 was introduced into the middle of the sn-2 chain at the 6- and 12-position for DPPC and DSPC, respectively. The C -13 and H-2 NMR spectra of each labeled lipid were studied as a functi on of temperature and CHOL concentration. The residual quadrupole spli tting in the H-2 NMR spectra, DELTAnu(Q perpendicular-to), was analyze d as a function of temperature and composition. For CHOL concentration s less than 30 mol %, a precipitous change in DELTAnu(Q perpendicular- to) occurs near the chain melting temperature of the phospholipid. Fur ther increases in CHOL concentration broaden the transition and shift the midpoint to higher temperature, indicating the presence of a new p hase at higher CHOL contents. Moreover, at a given temperature, DELTAn u(Q perpendicular-to) increases with increasing cholesterol content, w hich indicates a more ordered structure. The C-13 NMR spectra in the g el state consisted of a superposition of two components which can be a ttributed to both gel-like and fluid phospholipid domains in the bilay er. This two-component spectrum can be simulated quantitatively with a two-parameter chemical exchange model, which permits the fraction of each form and the exchange rate to be determined as a function of temp erature and composition. At high CHOL contents the line width of the f luid component broadens, suggesting an increase in the exchange rate b etween the domains. These results were interpreted in terms of a tempe rature composition diagram with one region L(beta'), two regions LG(I) and LG(II), and one liquid crystalline region L(alpha), with LG denot ing ''liquid-gel'' type phases. Liquid-gel phases correspond to phases with increased order in the hydrocarbon chains (in comparison to that of the pure PC bilayer in the L(alpha) phase) combined with fast limi t axial diffusion that averages the C-13 NMR spectrum to a ''fluidlike '' line. These phases are similar to those found in phosphatidylethano lamine/cholesterol (PE/CHOL) mixtures [Blume, A., & Griffin, R. G. (19 81) Biochemistry 24, 6230] and are in agreement with the results of Vi st and Davis [(1990) Biochemistry 29, 451].