Dynamic behavior analysis of ion transport through a bilayer lipid membrane by an electrochemical method combined with fluorometry.

To examine the transport of an ionic substance through a bilayer lipid membrane (BLM), an electrochemical method combined with fluorometry was proposed. In this method, the transport of a fluorescent ion through the BLM was detected both as the transmembrane current and the dynamic change of fluorescence intensity synchronizing scanning membrane potential. The fluorescence intensity was measured in the local area close to the planar BLM by utilizing a confocal fluorescence microscope. The electrochemical method combined with fluorometry makes it possible to analyze only the transport of a target fluorescent ion in distinction from the transport of other coexisting ions. With the proposed electrochemical method, the ion transport caused by both a hydrophobic fluorescent cation (rhodamine 6G+, R6G+) and a relatively hydrophobic anion (BF4-) was examined. The electrochemical method combined with fluorometry characterized the transmembrane current as the transport of R6G+. Membrane conductance for the R6G+ transport increased proportionally to the concentrations of R6G+ and BF4- distributed in the hydrocarbon medium of the BLM which were estimated by extraction experiments with liposomes. These results show that the distribution of a cation and an anion from the aqueous phase in the BLM predominantly controls the membrane conductance for ion transport through the BLM.

Distribution and Adsorption of Ionic Species into a Liposome Membrane and Their Dependence upon the Species and Concentration of a Coexisting Counterion.

The distribution of ions into a bilayer lipid membrane (BLM) and their adsorption on the BLM are investigated by extracting a hydrophobic cation, rhodamine 6G (R6G+), into a liposome through the dialysis membrane method. R6G+ distribution mainly depends upon the concentration of the coexisting anion and its species (Cl-, Br-, BF4-, ClO4-, and picrate). On the other hand, R6G+ adsorption on the BLM surface follows the Langmuir adsorption model and is independent of the coexisting anion in the aqueous phase. We propose an extraction model of ionic species into the BLM, to explain the dependence of extraction of ionic species upon the coexisting anion. In this model, an ion is distributed with a coexisting counterion into the BLM and then forms an ion pair in the BLM. Here, the ion adsorption equilibrium on the BLM surface is independent of the species and concentration of the coexisting counterion under the same ionic strength. On the basis of this model, we estimate the distribution constant of R6G+ and anion (KD), the ion-pair formation constant in the BLM (Kip), and the R6G+ adsorption constant on the BLM surface (Kad). Even for an ultrathin membrane system, such as a BLM, R6G+ is distributed with a coexisting counterion and the distribution equilibrium of the ionic species at the water-BLM interface is analyzable similar to that at the water-organic solvent interface.

Distribution of ion pairs into a bilayer lipid membrane and its effect on the ionic permeability.

This work reports the distribution constant of a target ion and a counter-ion between an aqueous phase and an artificial bilayer lipid membrane (BLM) and its influence to the ionic permeability through a BLM. A theoretical formula for ionic permeability through a BLM based on the distribution of the target ion and the counter-ion is also proposed and validated by analyzing the flux of a fluorescent cation [rhodamine 6G (R6G+)] through the BLM in the presence of counter-ions (X- = Br-, BF4-, and ClO4-). The transmembrane flux was evaluated by simultaneous measurement of the transmembrane current density and the transmembrane fluorescence intensity as a function of the membrane potential. The distribution constant of R6G+ and X- between the aqueous and BLM phases was determined by a liposome-extraction method. The measured ionic permeability exhibited non-linear dependent on the aqueous concentration of R6G+ or X-, but proportional to the concentration of R6G+ and X- inside the BLM evaluated from the distribution constant of R6G+ and X-. The proportionality demonstrates that the distribution of cations and anions between the aqueous and BLM phases dominates the flux of ion transport through the BLM. The proposed formula can express the dependence of the transmembrane current on the membrane potential and the concentrations of R6G+ and X- in the aqueous phase.