The passive transbilayer movementsflip-flopswas investigated on planar bilayer lipid membranes (BLMs), containing myristic, stearic, or linoleic long-chain fatty acids (FA). In response to a transbilayer pH gradient, a difference in the surface charges between inner and outer leaflets appeared. Because the BLM was formed from FA and neutral lipid, a surface potential difference was originated solely by a concentration difference of the initially equally distributed ionized FA. As revealed by

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... tential measurements, the corresponding surface potential difference ∆Φ s was at least twice the value expected from a titration of the FA alone. The additional surface charge was attributed to FA flip-flop induced by the transbilayer pH gradient. ∆Φ s was derived from capacitive current measurements carried out with a direct current (dc) bias and was corrected for changes of membrane dipole potential Φ d . Dual-wavelength ratiometric fluorescence measurements have shown that Φ d values of the pure DPhPC bilayers and BLMs containing 40 mol % FA differ by less than 6%. It is concluded that fast FA flip-flop is not restricted to membranes with high curvature. The role of pH gradient as an effective driving force for the regulation of FA uptake is discussed.