Phase separation in monolayers of pulmonary surfactant phospholipids at the air-water interface: Composition and structure

The phase behavior of monolayers containing the complete set of purified ph ospholipids (PPL) obtained from calf surfactant was investigated as a model for understanding the phase transitions that precede compression of pulmon ary surfactant to high surface pressure. During compression, both fluoresce nce microscopy and Brewster angle microscopy (BAM) distinguished domains th at separated from the surrounding film. Quantitative analysis of BAM graysc ales indicated optical thicknesses for the PPL domains that were similar to the liquid condensed phase for dipalmitoyl phosphatidylcholine (DPPC), the most abundant component of pulmonary surfactant, and higher and less varia ble with surface pressure than for the surrounding film. BAM also showed th e optical anisotropy that indicates long-range orientational order of tilte d lipid chains for the domains, but not for the surrounding film. Fluoresce nce microscopy shows that addition of DPPC to the PPL increased the area of the domains. At fixed surface pressures from 20-40 mN/m, the total area of each phase grew in proportion with the mol fraction of DPPC. This constant variation allowed analysis of the DPPC mol fraction in each phase, constru ction of a simple phase diagram, and calculation of the molecular area for each phase. Our results indicate that the phase surrounding the domains is more expanded and compressible, and contains reduced amounts of DPPC in add ition to the other phospholipids. The domains contain a mol fraction for DP PC of at least 96%.