Electron Spin Resonance Line-Shape Analysis of X-Doxylstearic Acid Spin Probes in Dihexadecyl Phosphate Vesicles and Effects of Cholesterol Addition

Electron spin resonance spectra of a series of x-doxylstearic acids (x = 5, 7, 10, 12, and 16), solubilized in anionic dihexadecyl phosphate vesicles containing 10, 30, and 50 mol % cholesterol, are analyzed by employing spectral line-shape simulation. The spectra from the vesicle phase are well reproduced by using the microscopic order-macroscopic disorder model of Freed and co-workers with Brownian rotational diffusion. The partial averaging of the magnetic interactions by local anisotropic motions in the vesicles is quantified mainly by the order parameter (S) and the rotational diffusion rate perpendicular to the alkyl chain (R(perpendicular-to)). The simulations show that the order parameter decreases with increasing x, suggesting a relatively extended conformation, unlike the U-shaped, bent conformation of the x-doxylstearic acid alkyl chain found previously in cationic dioctadecyldimethylammonium chloride vesicles. Furthemore, the headgroup region of the vesicle is less ordered in anionic dihexadecyl phosphate than in cationic dioctadecyldimethylammonium chloride vesicles. This result in liquid vesicle solutions corroborates the same x-doxylstearic conformations found in frozen vesicle solutions by electron spin echo modulation spectroscopy. The effect of cholesterol is found to decrease the observed order parameter for all doxyl positions. This effect is attributed to more water penetration into the vesicle interface arising from the intercalation of the large rigid cholesterol molecule. The effect of cholesterol addition is supported by using 3-doxyl-5alpha-cholestane, a spin probe which resembles cholesterol. It is found that the addition of cholesterol up to 50 mol % has little effect on the observed electron spin resonance spectrum of cholestane.