Effects of flunitrazepam on the Lalpha-H(II) phase transition of phosphatidylethanolamine using merocyanine 540 as a fluorescent indicator

Previous studies of our group demonstrated that flunitrazepam is a lipophilic drug capable of interacting with membranes through a partition equilibrium phenomenon. Its localization at the phospholipid polar head region could explain the decrease in the size of dipalmitoylphosphatidylcholine (dpPC) vesicles, through a mechanism that involves the increment in the relative volume of this region with a subsequent increase in the vesicle's surface curvature. In the present work, we investigated if flunitrazepam can affect the L(alpha)-H(II) phase transition of phosphatidylethanolamine through a similar mechanism. This study was approached by using merocyanine 540, a dye sensitive to the molecular packing of membrane lipids. A detailed analysis of merocyanine absorption and fluorescence emission and excitation spectra was performed. The results indicated that the fluorescence emitted came mainly from the monomeric form of merocyanine and that it resulted a good indicator of this phase transition, as was previously described. Flunitrazepam did not affect significantly the onset of the phase transition but showed a tendency to diminish the dye fluorescence emission intensity, which could involve a lower partition of merocyanine in the vesicles. Moreover, the results suggest that this drug produced a delay in the completeness of the phase transition and a decrement in the cooperativity of this phenomenon.