Deuterated Phospholipids as Raman Spectroscopic Probes of Membrane Structure: Dipalmitoylphosphatidylcholine-Dipalmitoylphosphatidylethanolamine Multilayers

Raman spectral data are reported for several conformation-sensitive spectral regions of dipalmitoylphos-phatidylethanolamine (DPPE) and for chain perdeuterated dipalmitoylphosphatidylcholine (DPL-d₆₂) alone, in 1:1 binary mixtures, and in 1:1:1 ternary combination with cholesterol. Raman melting curves for DPPE multilayers reveal the non-cooperative formation of 4-5 gauche rotamers per chain prior to the gel-liquid crystal transition at 66 °C. This phenomenon is also evident for DPL-d₆₂ multilayers. Use of d-62 as one component of a binary mixture allows the monitoring of conformational changes in each component individually. The phase separation of 1:1 DPL-d₆₂/DPPE mixtures shows onset and completion temperatures of about 44 and 60-65 °C, respectively, as monitored by variation in the line width of the C-D stretching vibrations of the DPL-d₆₂. The C-H stretching vibrations of the DPPE also respond to phase separation in the binary mixture. The I(2885)/(2850) intensity ratio shows maximal change close to the onset temperature and appearsto respond to both the formation of gauche rotamers anc changes in the lateral interactions of the DPPE chains. The frequency of the antisymmetric C-H stretching vibration n 2885 cm^-1 responds primarily to gauche rotamer format and exhibits a different temperature dependence than I(2885)/(2850) intensity parameter. The effect of deuteration on the phase separation is discussed. Addition of choleste to the binary mixture produces complex changes in the ph behavior. A broad phase separation region is still evident in ternary system, although the onset and completion tempe tures are each lower by about 15 °C. Cholesterol limits number of gauche rotamers that form during the phase se; ration from ~4 in binary DPL-d₆₂/DPPE to 1-2 in the pr ence of cholesterol. The presence of additional phases at low temperature is suggested in the ternary system.