We have previously shown that the ionophores lasalocid LAS), nigericin and monensin act as radical scavengers, inhibiting lipid peroxidation in mitochondria and in egg phosphatidylcholine (PC) liposomes containing negatively charged lipids and KCL as osmotic support. When tile osmotic support is sucrose, all ionophores failed to inhibit peroxidation. Here we analyse the kinetics of LAS membrane incorporation, a fundamental step for protection against lipid peroxidation. Despite the electrostatic repulsion, in the presence of KCI. the incorporation of tile anionic form (capable of complexing cations) is higher in negatively charged than in PC liposomes. Longer incubation times of LAS with PC liposomes increased ionophore incorporation, as attested by: i ) values of order parameter obtained from EPR spectra of lipid spin labels, ii) protection against lipid peroxidation (monitored by TBARS and conjugated diene), iii) LAS fluorescence and CD spectra. Sucrose prevents ionophore incorporation in both membrane systems. We conclude that the incorporation is characterized by a strong dependence on lipid charge, ionic compounds in the medium, and incubation time. Negatively charged membranes enhance the incorporation of LAS by increasing cation concentration at the interface, facilitating complexation, the limiting step of the process.119