A Highly K+‐Selective Two‐Photon Fluorescent Probe

A highly K⁺‐selective two‐photon fluorescent probe for the in vitro monitoring of physiological K⁺ levels in the range of 1–100 mM is reported. The two‐photon excited fluorescence (TPEF) probe shows a fluorescence enhancement (FE) by a factor of about three in the presence of 160 mM K⁺, independently of one‐photon (OP, 430 nm) or two‐photon (TP, 860 nm) excitation and comparable K⁺‐induced FEs in the presence of competitive Na⁺ ions. The estimated dissociation constant (K_d) values in Na⁺‐free solutions (K_d^OP=(28±5) mM and K_d^TP=(36±6) mM ) and in combined K⁺/Na⁺ solutions (K_d^OP=(38±8) mM and K_d^TP=(46±25) mM ) reflecting the high K⁺/Na⁺ selectivity of the fluorescent probe. The TP absorption cross‐section (σ_2PA) of the TPEF probe+160 mM K⁺ is 26 GM at 860 nm. Therefore, the TPEF probe is a suitable tool for the in vitro determination of K⁺.     

A Highly K+‐Selective Fluorescent Probe – Tuning the K+‐Complex Stability and the K+/Na+ Selectivity by Varying the Lariat‐Alkoxy Unit of a Phenylaza[18]crown‐6 Ionophore

A desirable goal is to synthesize easily accessible and highly K⁺/Na⁺‐selective fluoroionophores to monitor physiological K⁺ levels in vitro and in vivo. Therefore, highly K⁺/Na⁺‐selective ionophores have to be developed. Herein, we obtained in a sequence of only four synthetic steps a set of K⁺‐responsive fluorescent probes 4 , 5 and 6 . In a systematic study, we investigated the influence of the alkoxy substitution in ortho position of the aniline moiety in π‐conjugated aniline‐1,2,3‐triazole‐coumarin‐fluoroionophores 4 , 5 and 6 [R=MeO ( 4 ), EtO ( 5 ) and iPrO ( 6 )] towards the K⁺‐complex stability and K⁺/Na⁺ selectivity. The highest K⁺‐complex stability showed fluoroionophore 4 with a dissociation constant K_d of 19 mm , but the K_d value increases to 31 mm in combined K⁺/Na⁺ solutions, indicating a poor K⁺/Na⁺ selectivity. By contrast, 6 showed even in the presence of competitive Na⁺ ions equal K_d values (K_d^K+=45 mm and K_d^K+/Na+=45 mm ) and equal K⁺‐induced fluorescence enhancement factors (FEFs=2.3). Thus, the fluorescent probe 6 showed an outstanding K⁺/Na⁺ selectivity and is a suitable fluorescent tool to measure physiological K⁺ levels in the range of 10–80 mm in vitro. Further, the isopropoxy‐substituted N‐phenylaza[18]crown‐6 ionophore in 6 is a highly K⁺‐selective building block with a feasible synthetic route.     

A dual-fluorophore sensor approach for ratiometric fluorescence imaging of potassium in living cells

Potassium is the most abundant intracellular metal in the body, playing vital roles in regulating intracellular fluid volume, nutrient transport, and cell-to-cell communication through nerve and muscle contraction. On the other hand, aberrant alterations in K⁺ homeostasis contribute to a diverse array of diseases spanning cardiovascular and neurological disorders to diabetes to kidney disease to cancer. There is an unmet need for studies of K⁺ physiology and pathology owing to the large differences in intracellular versus extracellular K⁺ concentrations ([K⁺]_intra = 150 mM, [K⁺]_extra = 3–5 mM). With a relative dearth of methods to reliably measure dynamic changes in intracellular K⁺ in biological specimens that meet the dual challenges of low affinity and high selectivity for K⁺, particularly over Na⁺, currently available fluorescent K⁺ sensors are largely optimized with high-affinity receptors that are more amenable for extracellular K⁺ detection. We report the design, synthesis, and biological evaluation of Ratiometric Potassium Sensor 1 (RPS-1), a dual-fluorophore sensor that enables ratiometric fluorescence imaging of intracellular potassium in living systems. RPS-1 links a potassium-responsive fluorescent sensor fragment (PS525) with a low-affinity, high-selectivity crown ether receptor for K⁺ to a potassium-insensitive reference fluorophore (Coumarin 343) as an internal calibration standard through ester bonds. Upon intracellular delivery, esterase-directed cleavage splits these two dyes into separate fragments to enable ratiometric detection of K⁺. RPS-1 responds to K⁺ in aqueous buffer with high selectivity over competing metal ions and is sensitive to potassium ions at steady-state intracellular levels and can respond to decreases or increases from that basal set point. Moreover, RPS-1 was applied for comparative screening of K⁺ pools across a panel of different cancer cell lines, revealing elevations in basal intracellular K⁺ in metastatic breast cancer cell lines vs. normal breast cells. This work provides a unique chemical tool for the study of intracellular potassium dynamics and a starting point for the design of other ratiometric fluorescent sensors based on two-fluorophore approaches that do not rely on FRET or related energy transfer designs.

We report a dual-fluorophore approach for ratiometric fluorescent imaging of K⁺ levels in live cells. Intracellular esterases cleave RPS-1 to detach the K⁺-responsive fluorophore (PS525) from its internal standard (Coumarin 343).     

Oil-Water Partitioning of a Synthetic Tetracarboxylic Acid as a Function of pH

The oil-water partitioning of a synthetic tetraacid acting as a model compound for indigenous C₈₀-C₈₂ ARN acids has been studied as a function of pH, ionic strength and type of monovalent counterion. Experimental data obtained with ultraviolet-visible and HPLC/UV analyses have been fitted to thermodynamic models based on one, two or four dissociation steps to obtain o/w partition coefficients (K _wo ) of the fully protonated acid between chloroform and aqueous solutions, and its apparent acidity constant(s), pK _a. As the study is conducted above the CMC of the tetraacid, in general high apparent acidity constants were obtained in the range from 6 to 8 resulting from micellization equilibria. K _wo values were obtained in the range from 10^?3 to 10^?4, and decreasing with increasing salinity. At 50 mM K⁺, no conclusions could be made regarding the number of distinguishable dissociation steps, while at higher ionic strength (184 mM and 452 mM K⁺) and at 184 mM Na⁺ a model with two dissociation steps provided good fits to the experimental data. The first step was found to be given by a pK _a ≈ 6.6–6.8 and the second dissociation step at pK _a values ≈ 7.8–8.3. The two-step mechanism supports previous results obtained by potentiometric titrations. No significant difference in the o/w behavior was observed when changing the counterion from potassium to sodium. The main partitioning of the tetraacid in the aqueous phase occurred above pH 8, where the fully deprotonated acid was formed.     

Simple and sensitive fluorescence sensor for detection of potassium ion in the presence of high concentration of sodium ion using berberine–G-quadruplex complex as sensing element

In this work, a novel potassium ion (K⁺) sensor is presented using berberine–G-quadruplex complex as a fluorescent probe. This sensor is based on the K⁺that can induce the G-rich DNA to form G-quadruplex conformation. The G-quadruplex can bind berberine to form berberine–G-quadruplex complex, resulting in remarkable enhancement of fluorescence emission of the berberine–G-quadruplex system. In the presence of 800 mM sodium ion (Na⁺), the fluorescence of the berberine–G-quadruplex complex increased linearly with increasing K⁺ concentration in the range of 0.005–1.0 mM. The turn-on fluorescent assay is simple, inexpensive, and highly sensitive. We observed that Na⁺ in 10,000-fold molar excess does not interfere. The molecular mechanisms which produce enhanced fluorescence of berberine were discussed.     

A Highly K+‐Selective Phenylaza‐[18]crown‐6‐Lariat‐Ether‐Based Fluoroionophore and Its Application in the Sensing of K+ Ions with an Optical Sensor Film and in Cells

Herein, we report the synthesis of two phenylaza‐[18]crown‐6 lariat ethers with a coumarin fluorophore ( 1 and 2 ) and we reveal that compound 1 is an excellent probe for K⁺ ions under simulated physiological conditions. The presence of a 2‐methoxyethoxy lariat group at the ortho position of the anilino moiety is crucial to the substantially increased stability of compounds 1 and 2 over their lariat‐free phenylaza‐[18]crown‐6 ether analogues. Probe 1 shows a high K⁺/Na⁺ selectivity and a 2.5‐fold fluorescence enhancement was observed in the presence of 100 mM K⁺ ions. A fluorescent membrane sensor, which was prepared by incorporating probe 1 into a hydrogel, showed a fully reversible response, a response time of 150 s, and a signal change of 7.8 % per 1 mM K⁺ within the range 1–10 mM K⁺. The membrane was easily fabricated (only a single sensing layer on a solid polyester support), yet no leaching was observed. Moreover, compound 1 rapidly permeated into cells, was cytocompatible, and was suitable for the fluorescent imaging of K⁺ ions on both the extracellular and intracellular levels.     

ACE applied to the quantitative characterization of benzo‐18‐crown‐6‐ether binding with alkali metal ions in a methanol–water solvent system

ACE was applied to the quantitative evaluation of noncovalent binding interactions between benzo‐18‐crown‐6‐ether (B18C6) and several alkali metal ions, Li⁺, Na⁺, K⁺, Rb⁺ and Cs⁺, in a mixed binary solvent system, methanol–water (50/50 v/v). The apparent binding (stability) constants (K_b) of B18C6–alkali metal ion complexes in the hydro‐organic medium above were determined from the dependence of the effective electrophoretic mobility of B18C6 on the concentration of alkali metal ions in the BGE using a nonlinear regression analysis. Before regression analysis, the mobilities measured by ACE at ambient temperature and variable ionic strength of the BGE were corrected by a new procedure to the reference temperature, 25°C, and the constant ionic strength, 10 mM . In the 50% v/v methanol–water solvent system, like in pure methanol, B18C6 formed the strongest complex with potassium ion (log K_b=2.89±0.17), the weakest complex with cesium ion (log K_b=2.04±0.20), and no complexation was observed between B18C6 and the lithium ion. In the mixed methanol–water solvent system, the binding constants of the complexes above were found to be about two orders lower than in methanol and about one order higher than in water.     

Study of solvent effects on the stability constant and ionic mobility of the dibenzo‐18‐crown‐6 complex with potassium ion by affinity capillary electrophoresis

The effect of solvent on the strength of noncovalent interactions and ionic mobility of the dibenzo‐18‐crown‐6 complex with K⁺ in water/organic solvents was investigated by using affinity capillary electrophoresis. The proportion of organic solvent (methanol, ethanol, propan‐2‐ol, and acetonitrile) in the mixtures ranged from 0 to 100 vol.%. The stability constant, K_KL, and actual ionic mobility of the dibenzo‐18‐crown‐6‐K⁺ complex were determined by the nonlinear regression analysis of the dependence of the effective electrophoretic mobility of dibenzo‐18‐crown‐6 on the concentration of K⁺ (added as KCl) in the background electrolyte (25 mM lithium acetate, pH 5.5, in the above mixed hydro–organic solvents). Competitive interaction of the dibenzo‐18‐crown‐6 with Li⁺ was observed and quantified in mixtures containing more than 60 vol.% of the organic solvent. However, the stability constant of the dibenzo‐18‐crown‐6‐Li⁺ complex was in all cases lower than 0.5 % of K_KL. The log K_KL increased approximately linearly in the range 1.62–4.98 with the increasing molar fraction of organic solvent in the above mixed solvents and with similar slopes for all four organic solvents used in this study. The ionic mobilities of the dibenzo‐18‐crown‐6‐K⁺ complex were in the range (6.1–43.4) × 10^?9 m² V^?1 s^?1.     

Study of small chlorine‐doped potassium clusters by thermal ionization mass spectrometry

The theoretical calculations have predicted that nonmetal‐doped potassium clusters can be used in the synthesis of a new class of charge‐transfer salts which can be considered as potential building blocks for the assembly of novel nanostructured material. In this work, K_nCl (n = 2–6) and K_nCl_n?1 (n = 3 and 4) clusters were produced by vaporization of a solid potassium chloride salt in a thermal ionization mass spectrometry. The ionization energies (IEs) were measured, and found to be 3.64 ± 0.20 eV for K₂Cl, 3.67 ± 0.20 eV for K₃Cl, 3.62 ± 0.20 eV for K₄Cl, 3.57 ± 0.20 eV for K₅Cl, 3.69 ± 0.20 eV for K₆Cl, 3.71 ± 0.20 eV for K₃Cl₂ and 3.72 ± 0.20 eV for K₄Cl₃. The K_nCl⁺ (n = 3–6) clusters were detected for the first time in a cluster beam generated by the thermal ionization source of modified design. Also, this work is the first to report experimentally obtained values of IEs for K_nCl⁺ (n = 3–6) and K_nCl_n?1⁺ (n = 3 and 4) clusters. The ionization energies for K_nCl⁺ and K_nCl_n?1⁺ clusters are much lower than the 4.34 eV of the potassium atom; hence, these clusters should be classified as ‘superalkali’ species. Copyright © 2012 John Wiley & Sons, Ltd.     

The synthesis of bis(benzo-crown ether)s and their incorporation into potassium-selective PVC membrane electrodes

Five bis(benzo-15-crown-5) derivatives connected with different bridge chains were synthesized as neutral carriers in K⁺-selective electrodes. Potassium ion-selective PVC membrane electrodes based on these bis(crown ether)s were prepared using dibutyl phthalate (DBP) and dioctyl phthalate (DOP) as plasticizers of the PVC membrane. The selectivity coefficients (K _M ⁿ⁺:K _K+) for various alkali and alkaline-earth metal ions were measured. The electrodes based on the bis(crown ether)s are more selective for K⁺ than those based on monomeric crown ethers. The selectivity of one of the prepared potassium selective electrodes was higher than that of the electrode based on valinomycin and three of them were stable over a wide pH range.     

Non‐enzymatic Fructose Sensor Based on Co3O4 Thin Film

In this study, we report on the selective of fructose on Co₃O₄ thin film electrode surface. A facile chemical solution deposition technique was used to fabricate Co₃O₄ thin film on fluorine doped tin oxide, FTO, glass. Electrode characterization was done using XRD, HRTEM, SEM, AFM, and EIS. The constructed sensor exhibited two distinctive linear ranges (0.021–1.74 mM; 1.74–∼15 mM) covering a wide linear range of up to ∼15 mM at an applied potential of +0.6 V vs Ag/AgCl in 0.1 M NaOH solution. The sensor demonstrated high, reproducible and repeatable (R.S.D of <5 %) sensitivity of 495 (lower concentration range) & 53 (higher concentration range) μA cm⁻² mM⁻¹. The sensor produced a low detection limit of ∼1.7 μM (S/N =3). The electrode was characterised by a fast response time of <6 s and long term stability. The repeatability and stability of the electrode resulted from the chemical stability of Co₃O₄ thin film. The sensor was highly selective towards fructose compared to the presence of other key interferences i. e. AA, AC, UA. The ease of the electrode fabrication coupled with good electrochemical activity makes Co₃O₄ thin film, a promising candidate for non‐enzymatic fructose detection.     

Nafion‐modified Carbon Based Sensor for Soil Potassium Detection

In the present investigation, we report a voltammetric sensor based on Nafion modified screen printed carbon ink functionalized with 4‐aminobenzo‐18‐crown‐6 ether for the detection of soil potassium. The potassium is one of the major nutrient elements required by the plants and is critical for the crop productivity. The selective and sensitive assay is based on recording change in current signal of a positively charged redox probe, hexammineruthenium(III)chloride in the absence and presence of K⁺ ion in solution. The sensing interface is highly stable and reproducible for detection of K⁺ ion a under a wide range of concentration 1 to 500 ppm. The detection sensitivity over a wide concentration range was achieved by altering the concentration of redox species. Mechanistically, the recognition properties of 4‐aminobenzo‐18‐Crown‐6 covalently bound to the Nafion on carbon surface was found to be selective for potassium ion over sodium, ammonium and calcium ion. Potassium recognition through host‐guest binding of crown ether derivative and proposed simplicity of fabrication protocol makes the systems highly promising for real‐time applications in soil analysis.     

A Highly Selective Potassium Sensor for the Detection of Potassium in Living Tissues

The development of highly selective sensors for potassium is of great interest in biology. Two new hydrosoluble potassium sensors (Calix‐COU‐Alkyne and Calix‐COU‐Am) based on a calix[4]arene bis(crown‐6) and an extended coumarin were synthesized and characterized. The photophysical properties and complexation studies of these compounds have been investigated and show high molar extinction coefficients and high fluorescence quantum yields. Upon complexation with potassium in the millimolar concentration range, an increase of one‐ and two‐photon fluorescence emission is detected. A twofold fluorescence enhancement is observed upon excitation at λ=405 nm. The ligands present excellent selectivity for potassium in the presence of various competitive cations in water and in a physiological medium. The photophysical properties are not affected by the presence of a large amount of competing cations (Na⁺, Ca²⁺, Mg²⁺, etc.). Ex vivo measurements on mouse hippocampal slices show that Calix‐COU‐Alkyne accumulates extracellularly and does not alter the neuronal activity. Furthermore, the sensor can be utilized to monitor slow extracellular K⁺ increase induced by inhibition of K⁺ entry into the cells.     

Facilitated Transfer of Alkali and Alkaline‐Earth Metal Ions by a Calix[4]arene Derivative Across Water/1,2‐Dichloroethane Microinterface: Amperometric Detection of Ca2+

The host–guest complexation reactions between 5,11,17,23‐tetra‐tert‐butyl‐25,27‐diethoxycarbonylmethoxy‐26,28‐dimethoxy calix[4]arene (BDDC4) and alkali and alkaline‐earth metal ions were investigated by facilitated ion transfer processes across water/1,2‐dichloroethane microinterface by using steady‐state cyclic voltammetry and differential pulse voltammetry. The obtained facilitated transfers for Li⁺, Na⁺, K⁺, Rb⁺ and Ca²⁺ were evaluated under the different experimental conditions, at the excess concentrations of metal ions with respect to BDDC4 and vice versa. The association constants having 1 : 1 stoichiometry for Li⁺, Na⁺, K⁺ and Rb⁺ in 1,2‐DCE were determined. Also, we demonstrated that BDDC4 can play an important role for the development of highly selective chemical sensor for Ca²⁺ among alkaline‐metal ions in the concentration range of 0.1–1.0 mM in aqueous solution.     

[N‐(6‐Amino‐3,4‐di­hydro‐3‐methyl‐5‐nitroso‐4‐oxopyrimidin‐2‐yl)glycylglycinato]aquapotassium,a three‐dimensional coordination polymer

In the title compound, polymeric potassium N‐(6‐amino‐3,4‐di­hydro‐3‐methyl‐5‐nitro­so‐4‐oxopyrimidin‐2‐yl)­glycyl­gly­cinate hydrate, (K⁺·C₉H₁₁N₆O₅^?·H₂O)_n, the hexacoordinate K⁺ cation is linked to five different anions as well as to the water mol­ecule, with K—O distances in the range 2.617 (2)–2.850 (2) Å. Four of the O atoms in each anion coordinate to K centres, one of them acting as a bridging ligand, leading to the formation of nearly square centrosymmetric K₂O₂ rings. The structure is analysed in terms of (010) metal–ligand sheets linked by [010] chains of fused rings.     

Measurement of absolute rate data for the reaction of atomic potassium,K(42S½), with CH3F,C2H5F,C6H5F,CH3Br,C2H5Br,HCl and HBr by time-resolved atomic resonance absorption spectroscopy at λ = 404 nm (K(52PJ) ← K(42S½))

We present a kinetic study of atomic potassium in its electronic ground state, K(4²S_½), generated in the “single-shot mode” by pulsed irradiation at elevated temperatures and monitored by time-resolved atomic resonance absorption spectroscopy using the Rydberg doublet at λ = 404 nm (K[5²P_J]←K[4²S_½]). Profiles for the decay of atomic potassium in the presence of various halogenated reactants were recorded at different temperatures, yielding the following Arrhenius parameters (k_R = A exp(?E/RT), errors 1σ):

Vacant Manganese-Based Perovskite Fluorides@Reduced Graphene Oxides for Na-Ion Storage with Pseudocapacitive Conversion/Insertion Dual Mechanisms

Na-ion capacitors (NICs) and Na-based dual-ion batteries (Na-DIBs) have been considered to be promising alternatives to traditional lithium-ion batteries (LIBs) because of the abundance and low cost of the Na-ion, but their energy density, power density and life cycle are limited. Herein, dual-vacancy (including K⁺ and F⁻ vacancies) perovskite fluoride K_0.86MnF_2.69@reduced graphene oxide (rGO; recorded as Mn−G) as anode for NICs and Na-DIBs has been developed. The special conversion/intercalation dual Na-ion energy storage mechanism and pseudocapacitive dynamics are analyzed in detail. The Mn−G//AC NICs and Mn−G//KS6 Na-DIBs delivered a maximum energy density of 92.7 and 187.6 W h kg⁻¹, a maximum power density of 20.2 and 21.12 kW kg⁻¹, and long cycle performance of 61.3 and 68.4 % after 1000 cycles at 5 A g⁻¹, respectively. Moreover, Mn−G//AC NICs and Mn−G//KS6 Na-DIBs can work well over a wide range of temperatures (−20 to 40 °C). These results make it competitive in Na-ion storage applications with high energy/power density over a wide temperature range.     

Electrochemical Capacitive K+ EMIS Chemical Sensor Based on the Dibromoaza[7]helicene as an Ionophore for Potassium Ions Detection

A K⁺‐sensitive capacitive electrolyte‐membrane‐insulator‐semiconductor (EMIS) based on a novel dibromoaza[7]helicene ionophore has been developed. An ion‐sensitive membrane based on polyvinylchloride (PVC) doped with the ionophore was deposited on the Si₃N₄/SiO₂/Si‐p/Cu‐Al transducer. The properties of the K⁺‐EMIS chemical sensor were investigated by electrochemical impedance spectroscopy (EIS). All the developed devices upon being tested have shown good sensitivity and linearity responses within the range 10^?6 M to 10^?1 M of potassium activity, with good selectivity over a wide variety of other cations (Na⁺, Li⁺, Cu²⁺, Ca²⁺, and Mg²⁺). To our knowledge, this is the first time that a capacitive field‐effect sensor has been fabricated using helicene as a carrier for K⁺‐detection, combined with the structure: Si₃N₄/SiO₂/Si‐p/Cu‐Al as a transducer.     

Na+ Selective Fluorescent Tools Based on Fluorescence Intensity Enhancements,Lifetime Changes,and on a Ratiometric Response

Over the years, we developed highly selective fluorescent probes for K⁺ in water, which show K⁺-induced fluorescence intensity enhancements, lifetime changes, or a ratiometric behavior at two emission wavelengths (cf. Scheme 1, K1 – K4 ). In this paper, we introduce selective fluorescent probes for Na⁺ in water, which also show Na⁺ induced signal changes, which are analyzed by diverse fluorescence techniques. Initially, we synthesized the fluorescent probes 2 , 4 , 5 , 6 and 10 for a fluorescence analysis by intensity enhancements at one wavelength by varying the Na⁺ responsive ionophore unit and the fluorophore moiety to adjust different K_d values for an intra- or extracellular Na⁺ analysis. Thus, we found that 2 , 4 and 5 are Na⁺ selective fluorescent tools, which are able to measure physiologically important Na⁺ levels at wavelengths higher than 500 nm. Secondly, we developed the fluorescent probes 7 and 8 to analyze precise Na⁺ levels by fluorescence lifetime changes. Herein, only 8 (K_d=106 mm ) is a capable fluorescent tool to measure Na⁺ levels in blood samples by lifetime changes. Finally, the fluorescent probe 9 was designed to show a Na⁺ induced ratiometric fluorescence behavior at two emission wavelengths. As desired, 9 (K_d=78 mm ) showed a ratiometric fluorescence response towards Na⁺ ions and is a suitable tool to measure physiologically relevant Na⁺ levels by the intensity change of two emission wavelengths at 404 nm and 492 nm.     

Reactions of Aryl Benzoates with Potassium Aryloxides: Solvent Effects on Reaction Pathway and Kinetics

Temperature dependences of the relative reactivity of potassium aryloxides XC₆H₄O^?K⁺ toward 2,4‐dinitrophenyl benzoate in 50 mol% dimethylformamide (DMF)–50 mol% H₂O mixture have been studied using the competitive reactions technique. Correlation analyses of the relative rate constants k_X/k_H and differences in the activation parameters (ΔΔН^≠ and ΔΔS^≠) of the competitive reactions have revealed the existence of two isokinetic series of the reactions of 2,4‐dinitrophenyl benzoate with potassium aryloxides with electron‐donating substituent (EDS) and electron‐withdrawing substituent (EWS), respectively. We have investigated the effect of the substituent X on the activation parameters for each isokinetic series and concluded that the mechanism of the reactions of 2,4‐dinitrophenyl benzoate with potassium aryloxides XC₆H₄O^?K⁺ in 50 mol% DMF–50 mol% H₂O mixture is the same as in DMF. Analysis of the obtained data with using the method of two‐dimensional reaction coordinate diagram leads to the conclusion that the variation of the solvent from DMF to 50 mol% DMF–50 mol% H₂O mixture affects the reaction pathway. The rate constant k_X for the reaction of 3‐nitrophenyl benzoate with potassium 4‐methoxyphenoxide and the relative rate constants k_X/k_H for the reaction of 3‐nitrophenyl benzoate with potassium aryloxides XC₆H₄O^?K⁺ with EDS were measured in 50 mol% DMF–50 mol% H₂O mixtures at 25°C, and it has been shown that the addition of water to DMF does not change the mechanism but slows down these reactions.