Combining molecular modeling with experimental methodologies: mechanism of membrane permeation and accumulation of ofloxacin

The interaction between ofloxacin, as a model drug of the fluoroquinolone class, and biomembranes was examined as thepossible initial step in a transmembrane diffusion process. Dipalmitoylphosphatidylcholine was used for the preparation of biomembranemodels. The influence of environmental conditions and protonation on molecular physicochemical behavior, and henceon the membrane interaction, was investigated by differential scanning calorimetry (DSC). This technique has been shown to bevery effective in the interpretation of interactions of drug microspeciations with biomembranes. These findings suggest that theinteraction occurred owing to ionic and hydrophobic forces showing how the passage through the membrane is mainly favored inthe pH interval 6–7.4. It was demonstrated that a pH gradient through model membranes may be responsible for a poorly homogeneousdistribution of ofloxacin (or other related fluoroquinolones), which justifies the in vivo accumulation properties of thisdrug. DSC experiments, which are in agreement with computational data, also showed that the complexing capability of ofloxacinwith regard to Mg++ or Ca++ may govern the drug entrance into bacterial cells before the DNA Girase inhibition and couldensure the formation of hydrophobic and more fluid phospholipid domains on the surface of the model membrane. These regionsare more permeable with regard to various solutes, as well as ofloxacin, allowing a so-called ‘self-promoted entrance pathway’. Thecombination of experimental methodologies with computational data allowed a further rationalization of the results and openednew perspectives into the mechanism of action of ofloxacin, namely its interaction with lipid bilayers and drug–divalent cationcomplex formation, which might be extended to the entire fluoroquinolone class. Ofloxacin accumulation within Escherichia coliATCC 25922 was measured as a function of time. Also in this example, the environmental conditions influenced ofloxacin penetrationand accumulation. The in vitro experiments, reported here, show that a suitable balance of hydrophilic and hydrophobicfluoroquinolone properties needs to occur for there to be increased drug permeation.