We present a systematic study on how alkali metal salts, like NaCl and NaI, affect negatively charged phospholipid vesicles using a range of experimental methods. Our goal was to find out how chain saturation and cholesterol affect the interaction between the ions and the membrane. An isothermal titration calorimetry study on large unilamellar vesicles made from dimyristoyl phosphatidylcholine (DMPC) revealed that Na+ shows higher binding affinity to the gel phase at 15 °C compared to the fluid phase at 30 °C. Further, cations also show stronger affinity to the membrane in the fluid composed of saturated lipids than that of unsaturated lipids. The binding affinity of Na + with anionic vesicles prepared from a mixture of DMPC and DMPG was found to decrease significantly with increasing cholesterol as well as salt concentrations, as revealed by the zeta potential study. Besides the binding constant, the Gouy Chapman theory based on the electrostatic double layer shows that cholesterol reduces the surface charge density without altering the significant area per molecule. Further, the effect of counterions was investigated using fluorescence spectroscopy of an environment-sensitive lipophilic dye, nile red. Although cholesterol alters the emission properties of nile red significantly, there is no significant change in the presence of ions. This result suggests that anions do not bind significantly to anionic vesicles. The main striking feature of the ion-membrane interaction in the presence of cholesterol is that membranes with saturated lipids exhibit a completely opposite trend from membranes with unsaturated lipids.
Keywords: Cholesterol; Fluorescence spectroscopy; ITC; Membrane; NaI; Zeta potential.
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