The liquid-expanded to liquid-condensed phase transition of 1, 2-dipahmtoyl-sn-glycero-3-phosphocholine (DPPC) monolayers at the air-water (A/W) interface has been investigated in situ with external reflectance Fourier transform infrared spectroscopy. Infrared spectra have also been obtained for monolayer Films of 1, 2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC, a liquid-expanded film) and 1, 2-distearoyl-sn-glycero-3-phosphocholine (DSPC, a liquid-condensed film). All three phospholipid monolayer films were monitored in the conformation-sensitive C-H stretching region as a function of molecular area at the air-water interface. The measured frequencies of the symmetric and antisymmetric CH2stretching bands for the DPPC monolayer in the high molecular area, expanded phase (2853 and 2924 cm−1, respectively) are comparable to those of bulk DPPC multilayer dispersions above its main thermotropic phase transition and indicate a fluid and disordered conformation in the hydrocarbon chains. The measured frequencies for the low molecular area, condensed phase of DPPC (2849 and 2919 cm−1) indicate a rigid, mostly all-trans hydrocarbon chain conformation for the condensed phase monolayer, similar to the low-temperature, gel phase bulk DPPC dispersion. A continuous decrease in frequency with decreasing molecular area is observed throughout the transition region for the DPPC monolayer. These results demonstrate that the DPPC monolayer transition is (a) heterogeneous and biphasic in character, with coexistence of fluid and solid phases, and (b) involves a conformational change in the hydrocarbon chains of the monolayer as the average molecular area is decreased in the transition region. These trends are supported by the data for the DMPC and DSPC monolayer films. Molecular orientations relative to the substrate are also discussed in light of band intensities in both the parallel and perpendicular polarized spectra. © 1988, American Chemical Society. All rights reserved.