Relaxation dynamics of alkyl derivatives of fluorescein MitoFluo and C8-Fl in solutions with liposomes.

Abstract We present results of experimental and theoretical studies of excited state dynamics in two alkyl derivatives of fluorescein, MitoFluo and C8-Fl in solutions with liposomes. The liposomes DOPC and soybeanPC + 20% Cardiolipin (Azo-Cl), modelling cellular and inner mitochondrial membranes, respectively, were used in experiments. Both types of liposomes were shown to reduce significantly the fluorescence quantum yield as compared to that of pure fluorescein derivatives in solutions, while DOPC liposomes also caused a noticeable (ca 10 nm) red shift of fluorescence maximum. The study of fluorescence polarization decay has been carried out where important fluorescence parameters: polarization anisotropy, fluorescence lifetimes, and rotational diffusion times have been determined. It was shown that the isotropic fluorescence decay of C8-Fl in liposome containing solutions was single-exponential and the anisotropic decay was double-exponential for both types of lyposomes. In the case of MitoFluo both isotropic and anisotropic fluorescence decays were fitted satisfactory only with double-exponential functions. The interpretation of the experimental data obtained was supported by ab initio calculations of the structure and excitation properties of MitoFluo and C8-Fl in aqueous solution. The analysis of anisotropic fluorescence decay allowed for isolation of the contributions of fluorescein derivatives free in solution from those embedded in liposomes. Also, the experimental data suggest that MitoFluo interacts with liposomes more effectively than C8-Fl. Basing on the experimental and theoretical results obtained we conclude that free C8-Fl and MitoFluo molecules in solution were mostly in their dimer forms.