Real-Time Investigation of Lung Surfactant Respreading with Surface Vibrational Spectroscopy

The respreading of a lung surfactant monolayer at the air−water interface is investigated with broad bandwidth sum frequency generation (BBSFG) spectroscopy. The lung surfactant mixture contains chain perdeuterated dipalmitoylphosphatidylcholine (DPPC-d₆₂), palmitoyloleoylphosphatidylglycerol (POPG), palmitic acid (PA), and KL₄ (a 21-residue polypeptide analogue to the surfactant protein SP-B). DPPC-d₆₂ serves as a probe molecule for the spectroscopic investigation. The BBSFG spectra of DPPC-d₆₂ in the lung surfactant mixture are obtained in the C−D stretching region in real-time during film compression and expansion in a Langmuir trough. The BBSFG intensity of the CD₃ stretch peak from DPPC-d₆₂ terminal methyl groups is used as a measure of the interfacial density of DPPC-d₆₂ after careful consideration of orientation effects. For the first time, the interfacial loss of DPPC in a complex lung surfactant mixture is quantified. Spectroscopic results reveal that there is an 18% DPPC-d₆₂ interfacial loss during film respreading. However, the surface pressure−area isotherm measurements demonstrate that there is a rather large trough area reduction (37%) during film expansion. The relatively small interfacial loss of DPPC-d₆₂ and the rather large trough area reduction indicate that the respreading of DPPC and non-DPPC components in the lung surfactant is not uniform and a surface refinement process exists during film compression and expansion. This refinement process results in a DPPC-enriched monolayer with a significant depletion of non-DPPC components after film respreading. Implication for replacement surfactant design from this work is discussed.