Mobility and Molecular Orientation of Vitamin E in Liposomal Membranes as Determined by 19F NMR and Fluorescence Polarization Techniques

To elucidate the molecular orientation and mobility of α-tocopherol (vitamin E) in membranes, 19F nuclear magnetic resonance (NMR) was applied. In bilayer phosphatidyicholine (PC) liposomes, 19F NMR spin-spin relaxation times (T2) of 19F-labeled methyl groups on either the chromanol moiety or the isoprenoid side chain of atocopherol were significantly decreased compared to those for α-tocopherol in solution (CDCl3), suggesting that the vitamin E molecule exists in the lipid core of membranes and that its motional freedom is significantly restricted. The difference in T2 values between the solution state and PC liposomes was greatest for the 5a-CF3 on the chromanol moiety compared with other methyl groups on the isoprenoid side chain. Hence, it is thought that the chromanol moiety of α-tocopherol may fit tightly near the surface of the membrane because of its hydrophilicity. When Pr3+ was added to the liposonial suspension, the intensity of the NMR signal for 5a-CF3 changed faster than those of the other methyl groups. When the surface of the liposomal membranes was positively charged using stearyl amine, the mobility of the 5a-methyl group, expressed as the correlation time (τc), was quite similar to that in negatively charged membranes. These results indicate that the chromanol moiety of atocopherol may orient near the membrane surface, and that the hydroxyl group may be hydrogen bonded to the ester carbonyl of PC. For further confirmation of this conclusion, time-dependent changes in the fluidity of liposomes caused by the addition of Pr3+ were measured at the surface and inner region of liposomal membranes using a fluorescence polarization technique. The fluidity increased in the surface region and decreased in the inner region of liposomes containing α-tocopherol. These results show that when praseodymium ions may pass through the liposomal membrane, the hydrogen bonds between α-tocopherol and the ester carbonyl of PC may be broken, leading to the formation of an α-tocopherol-Pr3+ complex, thus causing the fluidity to increase. Based on the findings obtained, mobility and molecular orientation of vitamin E in membranes are proposed.