A study involving liposome-α-tocopherol organization and lipid peroxidation was carried out to contribute to the understanding of the correlation between the structure of the α-tocopherol carrier and the vitamin antioxidant activity in a mitochondrial membrane. Mitochondrial membranes were used as substrates for lipid peroxidation. α-tocopherol was incorporated in liposomes composed by 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) at different α-tocopherol concentrations: 1, 5, 10 and 20 mol% relative to DPPC. DPPC membrane packing was studied by Electron Spin Resonance (ESR) of spin labels incorporated into the liposomes and differential scanning calorimetry (DSC). Particle sizes were monitored by light scattering. As expected, ESR and DSC results revealed that α-tocopherol decreases DPPC rigidity when the bilayer is at the gel phase, and the gel-fluid transition is widened. Moreover, the presence of α-tocopherol in DPPC liposomes decreases mitochondrial peroxidation. Surprisingly, in the case of 10 and 20 mol% of α-tocopherol the decrement was found to be lower than with 1 and 5 mol%, with 1 mol% of α-tocopherol producing the best anti-peroxidant activity in mitochondrial membranes. In parallel, ESR and DSC data showed that with 1 mol% of α-tocopherol, at 30 oC, temperature at which the lipid peroxidation assay is performed, the DPPC bilayer is still quite packed, at gel state. At the same temperature, for concentrations of 5 mol% and above, DPPC enters in a broad gel-fluid transition, resulting in a less cooperative process. These findings could be related to the position of α-tocopherol active site nearby DPPC membrane surface. Accordingly, the activity was higher when DPPC membrane was more packed, in the gel phase; hence, the α-tocopherol active site would be more exposed, increasing the probability of α-tocopherol/free radical interaction; thus, decreasing mitochondrial membrane peroxidation. On the other hand, particle size analysis suggests that DPPC dispersions with higher α-tocopherol concentrations (above 5 mol%) are more aggregated than with 1 mol%. That, could also be relevant to the α-tocopherol antioxidant activity, as more α-tocopherol molecules could be exposed to the surface at lower α-tocopherol concentration. Hence, the present work shows that α-tocopherol anti-peroxidation activity in mitochondrial membranes is higher when the molecule is more diluted in DPPC membranes, the latter functioning major as a drug-carrier. That could be either related to the packing of DPPC vesicles and/or to their aggregation in the presence of higher α-tocopherol concentrations.

Liposomes, α-Tocopherol, Peroxidation, EPR, DSC, Carrier Structure, Light Scattering, Mitochondrial Membrane