Biophysical quantification of unitary solute and solvent permeabilities to enable translation to membrane science

Understanding the determinants of permeability and selectivity in biological channels serves two general pur-poses. It provides fundamental biophysical insights and allows the evaluation of new membrane proteins and artificial channel designs for the development of biomimetic separation membranes. This understanding relies on accurate ways to quantify unitary (single channel) solute and solvent permeabilities. However, the current research in biomimetic and bioinspired membranes and in protein biophysics tends to focus on relative per-meabilities. Further, many methods and approximations currently used result in erroneous permeability values which hinder and complicate the comparison between channels. Combined, these gaps in the available mea-surements obscure the biophysical insights and impede the development of novel high-performance channels. In this review, we summarize in vitro model membrane systems useful to functionally characterize artificial as well as biological channels. We also critically discuss biophysical techniques capable of extracting permeability values and membrane channel densities, which are fundamental parameters required to determine accurate unitary permeability values. Pertinent examples are provided, along with advantages, disadvantages, and potential improvements needed for specific techniques to acquire accurate unitary permeability values for a diverse set of small molecules, including water, ions, weak acids and bases, neutral molecules, and protons.