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Biomolecules 2019, 9(2), 63; https://doi.org/10.3390/biom9020063

The Effect of Buffers on Weak Acid Uptake by Vesicles

Institute of Biophysics, Johannes Kepler University Linz, Gruberstr. 40, 4020 Linz, Austria
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Received: 18 December 2018 / Revised: 7 February 2019 / Accepted: 8 February 2019 / Published: 13 February 2019
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Abstract

The assessment of weak acid membrane permeability (Pm) frequently involves large unilamellar vesicles. It relies on measurements of the intravesicular pH drop, ΔpHin, in response to a sudden augmentation of external acid concentration. However, ΔpHin may be primarily governed by non-instantaneous protonation and deprotonation reactions of (i) the acid itself, (ii) the buffer molecules, and (iii) the fluorescent pH reporter dye. Moreover, buffer concentration and acid gradient also serve as determinants of ΔpHin, as we show here. The uniexponential time constant (τ) of ΔpHin(t) is an invalid measure of Pm as Arrhenius plots of Pm and τ reveal different activation energies for acid influx. We calculate Pm by fitting a mathematical model to experimental stopped-flow traces. The model takes into account not only the time course of total internal buffer capacity but also (i) water self-dissociation, (ii) volume changes due to acid induced osmotic water flow, and (iii) the spontaneous membrane proton leak. It allows extracting a Pm of 30.8 ± 3.5 μm/s for formic acid for 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) vesicles. View Full-Text
Keywords: weak acid permeation; passive membrane permeability; membrane permeation; stopped flow; buffer; vesicles weak acid permeation; passive membrane permeability; membrane permeation; stopped flow; buffer; vesicles
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited (CC BY 4.0).

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Hannesschlaeger, C.; Barta, T.; Pechova, H.; Pohl, P. The Effect of Buffers on Weak Acid Uptake by Vesicles. Biomolecules 2019, 9, 63.

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