1. LAURDAN FLUORESCENCE AS AN INDICATOR OF SURFACE PRESSURE IN PULMONARY SURFACTANT FILMS M. Victoria Picardi*, Antonio Cruz, Jesús Pérez-Gil Dept. Bioquímica y Biología Molecular I, Fac. Biología, Universidad Complutense, Madrid, Spain (*picardiv@bbm1.ucm.es)  g DPPC  I B /I R I B -I R /I B +I R 0.28.90.52-0.32 0.413.10.76-0.14 0.612.60.68-0.19 0.821.30.80-0.11 1.027.40.92-0.04 1.227.51.370.16 1.429.61.350.15 1.632.12.170.37 1.838.03.380.54 2.438.81.620.24 INTRODUCTION Pulmonary surfactant, a lipid-protein complex secreted by alveolar type II cells, forms an interfacial film at the air-liquid interface of alveoli that reduces drastically the surface tension (close to 0 mN/m) avoiding lung collapse at the end of expiration. OBJECTIVES - Follow formation and dynamics of pulmonary surfactant films by spectroscopic techniques. RESULTS Evaluation of fluorescence Laurdan emission at different lipid densities in DPPC films using multiwell plates. Different aliquots of a DPPC solution 0,1 mg/ml in Chloroform/Methanol (2:1, vol/vol) were spread on top of a Tris 5 mM pH 7 subphase containing 150 mM NaCl deposited in different wells of a plate, to form films at different surface pressures. Afterwards, 2-5  L of a 0,07 mg/ml Laurdan solution in the same organic solvent mixture was spread on the surface of each well. Then, fluorescence emission spectra of Laurdan (excitation at 370 nm) was registered and surface pressure was measured in a surface balance. An Excitation Generalized Polarization function (Gp ex : I B -I R /I B +I R ) was selected to analyze all data in order to correlate Laurdan´s emission with the lateral packing of lipids in interfacial films. CONCLUSION Laurdan fluorescence is sensitive to changes in surface pressure in the range of 0-40 mN/m, suggesting the utility of this probe to monitorize lateral packing of lipids in interfacial films. Air space Capillar Endothelium Surfactant Interstitial layer Type I Pneumocyte Type II Pneumocyte Laurdan (6-Lauroyl-2-(N,N-dimethylamine) naphtalene) is a fluorescent probe used to analyse the structure of phospholipid membranes. When inserted into membranes, the fluorescence emission spectra of Laurdan is sensitive to the phase state of lipid molecules. Laurdan´s emission is blue-shifted in progressively packed lipid environments. In films with low lipid density the maximum emission of Laurdan occurs near 490 nm while in highly packed films the emission maximum shifts to around 440 nm, as a result of the reorientation of water dipoles around the excited- state probe dipole. I B : Fluorescence Intensity at 445 nm I R : Fluorescence Intensity at 490 nm  : Pressure 3005 20151025 Time 40 30 20 10 0 50 Pressure A good pulmonary surfactant must promote very rapid transfer of surface active phospholipids from bilayers at the aqueous phase into the air-liquid interface. This activity can be followed in  -t adsorption isotherms. - Use of Laurdan to evaluate the lateral packing of lipids in interfacial films. 440 490 Fluorescence Intensity Disordered Phase Ordered Phase Laurdan molecule 12.6 27.4 32.1 21.3 38.0 8.9 mN/m