Measuring Ca(2+) binding to short chain fatty acids and gluconate with a Ca(2+) electrode: role of the reference electrode.

Autor: Stumpff F; Institute for Veterinary Physiology, Department of Veterinary Medicine, Freie Universität Berlin, 14163 Berlin, Germany., McGuigan JA; Institute for Veterinary Physiology, Department of Veterinary Medicine, Freie Universität Berlin, 14163 Berlin, Germany. Electronic address: jasmcguigan@zedat.fu-berlin.de.
Jazyk: angličtina
Zdroj: Analytical biochemistry [Anal Biochem] 2014 Aug 15; Vol. 459, pp. 46-52. Date of Electronic Publication: 2014 May 15.
DOI: 10.1016/j.ab.2014.05.001
Abstrakt: Many organic anions bind free Ca(2+), the total concentration of which must be adjusted in experimental solutions. Because published values for the apparent dissociation constant (Kapp) describing the Ca(2+) affinity of short chain fatty acids (SCFAs) and gluconate are highly variable, Ca(2+) electrodes coupled to either a 3M KCl or a Na(+) selective electrode were used to redetermine Kapp. All solutions contained 130mM Na(+), whereas the concentration of the studied anion was varied from 15 to 120mM, replacing Cl(-) that was decreased concomitantly to maintain osmolarity. This induces changes in the liquid junction potential (LJP) at the 3M KCl reference electrode, leading to a systematic underestimation of Kapp if left uncorrected. Because the Na(+) concentration in all solutions was constant, a Na(+) electrode was used to directly measure the changes in the LJP at the 3 M KCl reference, which were under 5mV but twice those predicted by the Henderson equation. Determination of Kapp either after correction for these LJP changes or via direct reference to a Na(+) electrode showed that SCFAs do not bind Ca(2+) and that the Kapp for the binding of Ca(2+) to gluconate at pH 7.4, ionic strength 0.15M, and 23°C was 52.7mM.
(Copyright © 2014 Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
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