The First Online Capillary Electrophoresis-Microscale Thermophoresis (CE-MST) Method for the Analysis of Dynamic Equilibria-The Determination of the Acidity Constant of Fluorescein Isothiocyanate.

This article presents the first successful application of a capillary electrophoresis-microscale thermophoresis tandem technique (CE-MST) for determining the values of equilibrium constant, realized by connecting online the CE and MST instruments using a fused-silica capillary. The acid-base dissociation of fluorescein isothiocyanate, expressed by the acidity constant value (p), was used as a model. The measurement procedure consisted of introducing a mixture containing the analyte and a deliberately added interferent into the CE capillary, electrophoretic separation of the analyte from the interferent, the detection of the analyte with a CE-integrated detector, detection with a MST detector, and then stopping the flow temporarily by turning off the voltage source to conduct the thermophoretic measurement.

A sustainable approach for the stability study of psychotropic substances using vitreous humor and liver as alternative matrices.

The stability of psychotropic substances representing various drug groups important from the perspective of forensic chemistry, including benzodiazepines, antidepressants, carbamazepine, cocaine, and their selected metabolites, was investigated for 1 month in two alternative biological matrices, vitreous humor and liver homogenate. Three different thermal storage conditions (-20, 4, and 20 °C) were tested. Liquid chromatography-mass spectrometry (LC-MS) analysis was preceded by an effective solid-phase microextraction (SPME) procedure.

The Acid-Base/Deprotonation Equilibrium Can Be Studied with a MicroScale Thermophoresis (MST).

MicroScale thermophoresis (MST) is a rapidly developing bioanalytical technique used routinely for the examination of ligand-target affinity. It has never been used so far for the analysis of acid-base dissociation and the determination of p constant. This work is the-proof-of-concept of this new idea.

A Perspective of the Comprehensive and Objective Assessment of Analytical Methods Including the Greenness and Functionality Criteria: Application to the Determination of Zinc in Aqueous Samples.

The recently proposed concept of White Analytical Chemistry (WAC), referring to the Red-Green-Blue color model, combines ecological aspects (green) with functionality (red and blue criteria), presenting the complete method as "white". However, it is not easy to carry out an overall quantitative evaluation of the analytical method in line with the WAC idea in an objective manner. This paper outlines the perspective of the future development of such a possibility by attempting to answer selected questions about the evaluation process.

Comprehensive Assessment of Flow and Other Analytical Methods Dedicated to the Determination of Zinc in Water.

An original strategy to evaluate analytical procedures is proposed and applied to verify if the flow-based methods, generally favorable in terms of green chemistry, are competitive when their evaluation also relies on other criteria. To this end, eight methods for the determination of zinc in waters, including four flow-based ones, were compared and the Red-Green-Blue (RGB) model was exploited. This model takes into account several features related to the general quality of an analytical method, namely, its analytical efficiency, compliance with the green analytical chemistry, as well as practical and economic usefulness.

Differentiation of isomeric metabolites of carbamazepine based on acid-base properties; Experimental vs theoretical approach.

Metabolism of carbamazepine is complex and leads to the three isomeric derivatives whose occurrence is dependent on the type of sample material. Their unambiguous differentiation is overall important. In this work, the qualitative analysis of 2-hydroxycarbamazepine, 3-hydroxycrbamazepine and carbamazepine-10,11-epoxide was attempted for the first time using capillary zone electrophoresis, based on the models linking electrophoretic mobility with pK value determining the acidity of the hydroxyl groups.

Simultaneous quantification of food colorants and preservatives in sports drinks by the high performance liquid chromatography and capillary electrophoresis methods evaluated using the red-green-blue model.

Consumption of sports drinks is generally associated with a healthy lifestyle, despites a common addition of colorants and preservatives to these products. This work presents the novel methods based on high performance liquid chromatography (HPLC) and capillary electrophoresis (CE), aimed at the simultaneous quantification of both these groups of additives within a single separation. The results obtained show that colorants are generally added in low and rather safe amounts, whereas preservatives (benzoate and sorbate) are much more abundant, and likely pose a potential risk, especially for children.

Acidity constant of pH indicators in the supramolecular systems studied by two CE-based methods compared using the RGB additive color model.

Acid-base properties of methyl orange, bromocresol green, bromophenol blue, and bromothymol blue were thoroughly investigated in the past due to their application as colorimetric pH indicators. However, it is still unknown how these properties change upon the supramolecular host-guest interactions. Owing to the growing interest in using supramolecular host-guest interactions to reach expected modification of various physicochemical properties of guests, we decided to address this question in the present article.

Cyclodextrin-induced acidity modification of substituted cathinones studied by capillary electrophoresis supported by density functional theory calculations.

This paper shows that the acidity of substituted cathinones can change in a diversified and poorly predictable manner upon supramolecular interaction with cyclodextrins used as buffer additives in capillary electrophoresis. The direction and range of pK shifts may be noticeably different for the particular cyclodextrins and cathinones, suggesting a strict correlation with structure. The most interesting results were observed for 2-hydroxyethyl-β-cyclodextrin, which is capable for inducing the large negative and enantioselective apparent pK shifts for α-pyrrolidinovalerophenone and methylenedioxypyrovalerone, even much above -1.

Simultaneous enantioseparation of methcathinone and two isomeric methylmethcathinones using capillary electrophoresis assisted by 2-hydroxyethyl-β-cyclodextrin.

Methcathinone (ephedrone), 4-methylmethcathinone (mephedrone), and 3-methylmethcathinone (metaphedrone) are toxicologically-important cathinone derivatives used commonly as designer drugs. In this work we show the first method allowing to separate simultaneously all these molecules in a chiral medium, ensuring good resolution between all enantiomers. Eight cyclodextrins have been tested as potential chiral selectors, the best results were obtained with 2-hydroxyethyl-β-cyclodextrin, unreported so far for efficient separation of cathinones.

Flow variation as a factor determining repeatability of the internal standard-based qualitative and quantitative analyses by capillary electrophoresis.

The use of migration times and peak areas referred to another sample component - internal standard, brings many benefits in improving reliability of capillary electrophoresis. However, it is quite commonly overlooked that despite relative migration time and peak area ratio are more stable than the absolute values upon alteration in the flow rate, some shift should always be expected. The present work offers a new look at this analytically-important issue.

Thermodynamics of acid-base dissociation of several cathinones and 1-phenylethylamine, studied by an accurate capillary electrophoresis method free from the Joule heating impact.

Capillary electrophoresis is often used to the determination of the acid-base dissociation/deprotonation constant (pK), and the more advanced thermodynamic quantities describing this process (ΔH°, -TΔS°). Remarkably, it is commonly overlooked that due to insufficient dissipation of Joule heating the accuracy of parameters determined using a standard approach may be questionable. In this work we show an effective method allowing to enhance reliability of these parameters, and to estimate the magnitude of errors.

Acidity of substituted cathinones studied by capillary electrophoresis using the standard and fast alternative approaches.

Cathinone derivatives are notorious but still weakly characterized molecules, known mainly as components of the designer and illicit drugs. The knowledge on their acidity is scarce and incomplete, therefore, we decided to determine the pK values for six of them: 2-methylmethcathinone, 3-methylmethcathinone, 4-methylmethcathinone, α-pyrrolidinovalerophenone, methylenedioxypyrovalerone and ephedrone. For that purpose we employed capillary electrophoresis, which is known for its accurateness in comparison to other analytical techniques.

Improving repeatability of capillary electrophoresis-a critical comparison of ten different capillary inner surfaces and three criteria of peak identification.

A poor repeatability of migration times caused by the fluctuations of electroosmotic flow (EOF) is an inherent weakness of capillary electrophoresis. Most researchers endeavor to prevent this problem using relative migration times or various capillary coatings which are expensive and not easy in comparison. Herein, we present an original approach to this problem: we apply a model sample designed to induce significant EOF instability, in order to critically compare ten capillary types with different physicochemical characteristics.

Origin of Remarkably Different Acidity of Hydroxycoumarins-Joint Experimental and Theoretical Studies.

In the present work the origin of highly varied acidity of hydroxycoumarins (pK values) has been for the first time investigated by joint experimental and computational studies. The structurally simple regio-isomers differing in the location of hydroxyl group, 3-hydroxycoumarin (3-HC), 4-hydroxycoumarin (4-HC), 6-hydroxycoumarin (6-HC), 7-hydroxycoumarin (7-HC), as well as 4,7-dihydroxycoumarin (4,7-HC) and the larger 4-hydroxycoumarin-based derivatives: warfarin (WAR), 7-hydroxywarfarin (W7), coumatetralyl (CT), and 10-hydroxywarfarin (W10), have been compared in terms of enthalpy-entropy relationships accounting for the observed pK values. We have revealed that in the case of large molecules the acidic proton is stabilized by the following noncovalent interactions OH···O (WAR and W7), OH···π (CT), and OH···OH···O (W10), this effect leads to a compensatory enthalpy-entropy relation and yields a moderate pK increase.

Enhancing effectiveness of capillary electrophoresis as an analytical tool in the supramolecular acidity modification.

A strategic modification of acidity (pK values) by the non-covalent host-guest interactions is one of the most promising concepts in current supramolecular chemistry. This work is aimed at enhancing the effectiveness of capillary electrophoresis (CE) in determination of pK shifts caused by such interactions and their thermal dependencies crucial in a deep thermodynamic description. We show how to (i) minimize the systematic errors related to Joule heating, (ii) minimize the influence of a voltage ramp time, (iii) speed up pK shift identification and estimation, (iv) interpret thermal effects related to two overlapped dynamic equilibria, and (v) determine pK shifts by an alternative spectrophotometric method (CE-DAD).

Minimizing the impact of Joule heating as a prerequisite for the reliable analysis of metal-protein complexes by capillary electrophoresis.

Herein we report on a drastic release of metal ions from the Fe-bound transferrin, and Fe- or Mn-bound lactoferrin, observed upon the increase in the separation voltage during CE-based analysis. To verify whether this process is caused directly by electric field, we developed an Isothermal Voltage Increase approach (IVI), which is the extension of methods reported by Krylov et al. IVI ensures isothermal conditions while increasing separation voltage by a hydrodynamic pushing of the injected sample to the actively cooled capillary section, combined with a rationale choice of cooling temperature, dependent on the value of current.

Seven Approaches to Elimination of the Inherent Systematic Errors in Determination of Electrophoretic Mobility by Capillary Electrophoresis.

Electrophoretic mobility is a basic parameter that describes the electromigration of an ionized particle, which is used in many fields of analytical and physicochemical science. Its determination by capillary electrophoresis (CE), using a routine method, is intrinsically affected by the generation of Joule heating, entailing a drop in viscosity and possible alteration of the degree of ionization, and also by other commonly overlooked effects: axial electric field distortion and voltage ramping. The objective of this work was to provide the first theoretical overview and experimental comparison of all accessible methods that could be used to prevent these sources of inaccuracy.

Capillary coating as an important factor in optimization of the off-line and on-line MEKC assays of the highly hydrophobic enzyme chlorophyllase.

The choice between bare and coated capillaries is a key decision in the development and use of any methods based on capillary electrophoresis. In this work several permanently and dynamically coated capillaries were successfully implemented in a previously developed micellar electrokinetic chromatography (MEKC) assay of the plant membrane enzyme chlorophyllase. The results obtained demonstrate the rationale behind the use of capillary coating, which is crucial for successful optimization of both the off-line mode and the on-line/electrophoretically mediated microanalysis assay mode.

Cyclodextrin-assisted enantioseparation of warfarin and 10-hydroxywarfarin by capillary electrophoresis studied from the analytical and thermodynamic points of view.

In this work cyclodextrin-assisted enantioseparation of warfarin and 10-hydroxywarfarin by CE has been studied from the analytical and thermodynamic points of view. The role of cyclodextrin concentration and temperature has been analyzed in reference to three different analytical parameters, corresponding to selectivity, resolution and resolution/analysis time ratio. The optimal conditions for enantioseparation have been found, they have been selected on the basis of critical difference in electrophoretic mobility and possibly short analysis time.

Determination of acid dissociation constant of 20 coumarin derivatives by capillary electrophoresis using the amine capillary and two different methodologies.

In this work capillary electrophoresis has been used to determine acid dissociation constant of 20 structurally diverse coumarin derivatives. For a majority of compounds pKa value has been determined for the first time. The obtained values vary between 4.

A comparative study of various physicochemically modified capillaries used in CE technique for the three distinct analytical purposes.

The five different commercially offered capillaries, bare silica, one dynamically and three permanently coated, have been tested with respect to the three distinct bioanalytical issues: (i) achiral and chiral separation of small mass molecules, warfarin and its six isomeric hydroxy-derivatives; (ii) ultraselective separation of transferrin and albumin including differentiation between the diferric, two monoferric and iron-free forms of transferrin; and (iii) pioneering identification of albumin-induced shifts of acid dissociation constant by using CE instrumentation, revealed for warfarin and six hydroxywarfarins. As a result all of these pharmacologically and biochemically-relevant purposes have been reached, but using different capillaries. The dynamically coated silica capillary has allowed for the first time to separate warfarin from its six main hydroxy-metabolites by CE with high resolution.

A simple method for assessment and minimization of errors in determination of electrophoretic or electroosmotic mobilities and velocities associated with the axial electric field distortion.

It is commonly accepted that the modern CE instruments equipped with efficient cooling system enable accurate determination of electrophoretic or electroosmotic mobilities. It is also often assumed that velocity of migration in a given buffer is constant throughout the capillary length. It is simultaneously neglected that the noncooled parts of capillary produce extensive Joule heating leading to an axial electric field distortion, which contributes to a difference between the effective and nominal electric field potentials and between velocities in the cooled and noncooled parts of capillary.