Hydrophobic barrier in liposomes studied by FT-ESEEM

An improvement of the electron spin echo envelope modulation technique (ESEEM) for studying the hydrophobic barrier in lipid membranes was proposed. Water penetration depth into the lipid bilayer composed of egg-phosphatidylcholine and dicetylphosphate was studied. For this purpose the stearic acid spin probes with nitroxide moiety at different positions on the acyl chain were dissolved in the bilayer of liposomes prepared in buffer solution with D2O. The ESEEM method is based on the deuterium effect to the 14N-hyperfine splitting constant of the spin probe. For the analysis of the spectra we suggested the frequency domain approach (Fourier transform-ESEEM) instead of the time domain. In order to test this approach, the influence of cholesterol and octanol on the water penetration depth into liposomes was measured. FT-ESEEM appeared to be a more sensitive method for the quantitative measurements of the water penetration depth than the time domain. This approach can become one of the most effective methods for studying the influence of surfactants and other biologically active molecules on water penetration depth into membranes.