A new 19F NMR indicator for intracellular sodium

Abstract

The effect of increased benzo substitution on the cation-binding strength of diaza-18-crown-6 has been investigated. The planarity constraint imposed by increased substitution reduces the effective cavity size thus giving rise to an increased selectivity for sodium over potassium, magnesium and calcium that is sufficient to provide a class of indicators suitable for measuring intracellular free sodium. The cation binding parameters and the associated spectral changes of a selection of compounds with varied aryl fluorination patterns have shown that there is very little electronic interaction between the aromatic rings and the bound cation through the heteroatoms of the crown ether. Only small adjustments to the cation affinity can therefore be achieved via alteration of the aryl substitution patterns. Aryl substitution adjacent to the hetero atoms gives a reduction in the cation affinity of the tribenzo-crown ether, similar to that found on fluorination of the calcium chelator BAPTA ortho to the oxyethylene bridge. These observations have led to the preparation of a new indicator for intracellular sodium that we have called FCrown-1. The new indicator has a dissociation constant for sodium of 11 mmol dm" and a selectivity for sodium over potassium of 173-fold. The sodium concentration is reported by the chemical shift (fast exchange) of one of the two types of fluorine present with a maximum sodium-induced shift of 4.6 ppm downfield. The resonance arising from the other fluorine substituents is insensitive to sodium and acts as an internal chemical shift reference at 8.8 to 13.4 ppm upfield from the reporting signal.Paper 2/05554F

Synthesis and properties of a potential extracellular fluorescent probe for potassium

“…l 6 The fluorescent properties of the new probes were examined under the conditions used in our earlier studies and followed the procedure described by Minta and Tsien.' Vilsmeier-Haack formylation, in the manner described above, gave the bisaldehyde 17.…”

A method for multiparametric statistical quantification of the heterogeneity of free Na+concentration by19F MR spectroscopy: Proof of principlein silicoandin vitro

“…The relationship between the measured chemical shift, δ mea , and [Na + ] is given by the following equations, in strict analogy with equations previously used for [Ca 2+ ] and [Mg 2+ ]: with , dissociation constant of complex between Na + and the ligand, FCrown‐1; [Cro] f , concentration of free (uncomplexed) FCrown‐1; [FCro] tot , total concentration of FCrown‐1 (free and complexed with Na + ); δ Cro , meta ‐ 19 F chemical shift for free (uncomplexed) FCrown‐1; δ NaCro , meta ‐ 19 F chemical shift for FCrown‐1 complexed with Na + . Experimentally determined , δ Cro and δ NaCro values have been published previously . They may vary slightly between different biological tissues as a function of intracellular parameters such as pH and ionic strength, and are, therefore, dependent on suitable calibration on the basis of solutions mimicking the pH and ionic strength of the tissue in question.…”

“…These spectra exhibit two resonances stemming from fluorine in meta and ortho positions in relation to the oxoethylene bridge. While the 19 F chemical shift representing the meta ‐fluorine increases by up to 4.6 ppm (from 8.8 to 13.4 ppm) with increasing [Na + ], the resonance from ortho ‐fluorine is independent of [Na + ] and, therefore, is used as a fixed internal standard for chemical‐shift referencing …”

“…The in silico modeling parameters, ie (i) the 19 F NMR chemical shift of meta ‐FCrown‐1 as a function of [Na + ], (ii) the 19 F NMR linewidth of meta ‐FCrown‐1, and (iii) the Na + /FCrown‐1 dissociation constant, are adopted from previously published experimental results . In addition, we subject in vitro spectra to our algorithms designed to extract statistical [Na + ] descriptors.…”

New frontiers and developing applications in 19F NMR

“…Fluorinated NMR reporters have also been demonstrated for [Mg 2+ ] and [Na + ] [276–280] and tested in biological systems including multiparametric assessment of Ca 2+ , Mg 2+ and pH in perfused ferret hearts [276]. Many other metal ion binding fluorinated ligands have been reported with rings optimized to accommodate the metal ion of interest, notably series of cyclams and cryptands, though these do not appear to have been used in vivo [281, 282] (Table 1).…”