Solvent Relaxation in Biomembranes

Fluorescence spectroscopy has considerably contributed to the present picture of the structure and function of biomembranes consisting predominantly of phospholipids. Among the fluorescence techniques employed such as quenching [1], energy transfer [2, 3], lifetime (distributions) [4–6], and excimer formation [7, 8], the determination of fluorescence anisotropy [4] has certainly been the dominating fluorescence method in studies of biological and model membranes. Fluorescence polarization studies, however, exhibit some major limitations: the experimentally determined steady-state and time-resolved anisotropics characterize the motional restrictions of the ‘reporter’ molecule itself and give therefore only indirect information about the dye environment, with the consequence that, if the probe is bound covalently to the lipid [e.g., l-(4-tri-methylammoniumphenyl)-6-phenyl-l,3,5-hexatriene; TMA-DPH], this attachment may dominate the recorded depolarization behavior. The membrane order parameters obtained from freely mobile probes like l,6-diphenyl-l,3,5-hexatriene (DPH) result from a broad distribution of localization within the hydrophobic interior, the detailed characterization of which reveals inherent ambiguities [9]. Moreover, the anisotropy technique is limited to the characterization of the hydrophobic bilayer interior, since the anisotropy of head-group labelled phospholipids appears to be rather insensitive to environmental changes [4]. These limitations do not restrict the application of the solvent relaxation method, the principles of which and their applications in membrane studies will be presented in this review.