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  • Title: Origin of laurdan sensitivity to the vesicle-to-micelle transition of phospholipid-octylglucoside system: a time-resolved fluorescence study.
    Author: Viard M, Gallay J, Vincent M, Paternostre M.
    Journal: Biophys J; 2001 Jan; 80(1):347-59. PubMed ID: 11159407.
    Abstract:
    The fluorescent probe laurdan has been shown to be sensitive to the vesicle-to-micelle transition of phosphatidylcholine/octylglucoside (M. Paternostre, O. Meyer, C. Grabielle-Madelmont, S. Lesieur, and, Biophys. J. 69:2476-2488). On the other hand, a study on the photophysics of laurdan in organic solvents has shown that the complex de-excitation pathway of the probe can be described by two successive processes, i.e., an intramolecular charge transfer followed by dielectric relaxation of the solvent if polar. These two excited-state reactions lead to three emitting states, i.e., a locally excited state, a charge transfer state, and a solvent relaxed state (M. Viard, J. Gallay, M. Vincent, B. Robert and, Biophys. J. 73:2221-2234). Experiments have been performed using time-resolved fluorescence on the probe inserted in amphiphile aggregates (mixed liposomes, mixed micelles) different in detergent-to-lipid ratios. The results have been compared with those obtained for laurdan inserted in dipalmitoyl phosphatidylcholine liposomes in the gel and in the fluid lamellar phase. Except for laurdan in dipalmitoyl phosphatidylcholine liposomes in the gel lamellar phase, the red part of the emission spectra originates from the de-excitation of the relaxed excited state of laurdan, indicating that indeed the dielectric relaxation process is an important phenomena in the ground-state return pathway of this probe. On the other hand, the maximization entropy method (MEM) analysis of the fluorescence decay recorded in the blue part of the emission spectra indicates that the dielectric relaxation is not the only reaction occurring to the excited state of laurdan. Moreover, the analysis of the fluorescence decays of laurdan inserted in gel lamellar dipalmitoylphosphatidylcholine (DPPC) liposomes indicates excited-state reactions, although dielectric relaxation is impossible. These results are in agreement with the de-excitation pathway determined from laurdan behavior in organic solvent even if, in most of the aggregates studied in this work, the major phenomenon is the dielectric relaxation of the solvent. All along the vesicle-to-micelle transition, we have observed that the lifetime of the relaxed excited state of laurdan continuously decreases probably due to a dynamic quenching process by water molecules. On the other hand, the time constant of the dielectric relaxation process remains almost unchanged in the lamellar part of the transition but abruptly decreases as soon as the first mixed micelle is formed. This decrease is continuous all over the rest of the transition even if it is more pronounced in the mixed liposomes' and mixed micelles' coexistence. The increase of the octylglucoside-to-lipid ratio of the mixed micelles via the change of the size and the shape of the aggregates may facilitate the penetration and the mobility of water molecules. Therefore, during the vesicle-to-micelle transition, laurdan probes the evolution of both the amphiphile packing in the aggregates and the increase of the interface polarity. This study finally shows that the detergent-to-lipid ratio of the mixed micelles is an important parameter to control to limit the penetration and the mobility of water within the amphiphile aggregates and that laurdan is a nice tool to monitor this phenomenon.
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