Erzsébet Varga

Enantiomeric separation of antimalarial drugs by capillary electrophoresis using neutral and negatively charged cyclodextrins

Capillary electrophoresis (CE) methods for chiral resolution of five antimalarial drugs (primaquine, tafenoquine, mefloquine, chloroquine and quinacrine) were developed by using a wide selection of neutral and anionic cyclodextrin (CD) derivatives. The use of sulfobutyl-β-CD and carboxymethyl-β-CD (CMBCD) resulted in good resolution of quinacrine and tafenoquine, respectively. New results are presented for resolutions of chloroquine and mefloquine. Application of carboxyalkyl- and sulfobutyl-CD derivatives provided improved resolution for primaquine. The impurity in primaquine sample detected by CE was identified as quinocide by MS and NMR. CMBCD provided not only the best separation of primaquine from quinocide but also the simultaneous complete resolution of both compounds.

Separation of vinca alkaloid enantiomers by capillary electrophoresis applying cyclodextrin derivatives and characterization of cyclodextrin complexes by nuclear magnetic resonance spectroscopy

In this work, the enantiomeric separation of three vinca alkaloid enantiomers (vincamine, vinpocetine and vincadifformine) has been investigated in an aqueous capillary electrophoresis (CE) system using cyclodextrins (CDs). The investigated CDs were the native alpha-, beta-, and gamma-CDs and their hydroxypropylated, randomly methylated, carboxymethylated and sulfobutylated derivatives. The first part of this study consisted of the determination of the apparent averaged complex stability constants with the selected CDs. Several parameters, such as the nature and the concentration of the CD, were studied and were found to have a significant effect on the enantiomeric resolution for all studied compounds. All three vinca alkaloids were successfully enantioseparated with CDs where different migration orders were observed in case of several CDs depending on the cavity size or substituent of the host. Chiral separation and determination of the stability constants were also performed with NMR spectroscopy which confirmed the CE results. Averaged stoichiometries of the complexes were determined using the Job plot method resulting in a 1:1 complex irrespective of the alkaloid enantiomers or cyclodextrin derivative. The structures of the inclusion complexes were elucidated using 2D ROESY NMR spectroscopy. On the basis of NMR results reversal of enantiomer migration order was clarified in various cases.

Characterization of aspartame-cyclodextrin complexation

The inclusion complex formation of aspartame (guest) and various cyclodextrins (host) were examined using 1H NMR titration and capillary electrophoresis. Initially the protonation constants of aspartame were determined by NMR-pH titration with in situ pH measurement to yield log K1=7.83 and log K2=2.96. Based on these values the stability of the complexes formed by aspartame and 21 different cyclodextrins (CDs) were studied at pH 2.5, pH 5.2 and pH 9.0 values where aspartame exists predominantly in monocationic, zwitterionic and monoanionic form, respectively. The host cyclodextrin derivatives differed in various sidechains, degree of substitution, charge and purity so that the effect of these properties could be examined systematically. Concerning size, the seven-membered beta-cyclodextrin and its derivatives have been found to be the most suitable host molecules for complexation. Highest stability was observed for the acetylated derivative with a degree of substitution of 7. The purity of the CD enhanced the complexation while the degree of substitution did not provide obvious consequences. Finally, geometric aspects of the inclusion complex were assessed by 2D ROESY NMR and molecular modelling which proved that the guests aromatic ring enters the wider end of the host cavity.

Separation of cis-β-lactam enantiomers by capillary electrophoresis using cyclodextrin derivatives

Chiral separation of 19 pairs of cis-beta-lactam (BL) stereoisomers of pharmacological importance was examined by capillary electrophoresis using cyclodextrin (CD) derivatives. In order to select the most effective conditions separating the highest number of stereoisomers of BLs, single carboxymethyl alpha-, beta- and gamma-, as well as sulfobutyl beta-CD derivatives were applied. Additionally, carboxymethyl and sulfobutyl beta-CD derivatives complemented with neutral beta-CD derivatives as dual CD systems were tested. Both the composition and concentration of applied selectors and the pH of background electrolyte were selected. In single systems the structural characteristics of BLs and the complex forming affinity were correlated. Most BLs provided optimal complexation with beta-CD derivatives. In conclusion, the efficiency of combining sulfobutyl-beta-CD and permethylated beta-CD was superior to other single and dual CD systems applied. This method successfully separated each pair of stereoisomers investigated.

Single-isomer carboxymethyl-γ-cyclodextrin as chiral resolving agent for capillary electrophoresis

Herein we report on the synthesis, characterization and the novel capillary electrophoretic use of octakis-(2,3-di-O-methyl-6-O-carboxymethyl)-γ-cyclodextrin sodium salt (ODMCM). ODMCM is the first single-isomer carboxymethyl-γ-cyclodextrin that is fully methylated on its secondary side and carries ionizable carboxymethyl functions on its primary side. ODMCM was prepared with high isomeric purity through a four-step synthetic procedure. The purity of each intermediate was characterized by appropriate chromatographic methods, while the isomeric purity of the carboxymethylated product was determined by an HPLC method using a CD-Screen-IEC column and by a capillary electrophoretic method using indirect UV detection, as well. The structural identification of the ODMCM was carried out by 1D, 2D NMR spectroscopy and ESI-MS. The acid-base characterization of the chiral selector was carried out by 1H NMR-pH titration. The chiral separation ability of the synthesized selector was studied by chiral capillary electrophoresis. ODMCM was used as a background electrolyte additive to separate enantiomers of representative pharmacologically significant model molecules such as propranolol, citalopram, ketamine, tapentadol and dapoxetine. The effects of the selector concentration and the pH of the background electrolyte on the enantiorecognition properties were investigated. 1H NMR spectroscopy was further applied to get deeper insight of the host-guest inclusion complex formation. The pH-dependent enantioselectivity of this new single-isomer chiral selector was demonstrated by chiral capillary electrophoresis and 1H NMR spectroscopy.

Comparative evaluation of the chiral recognition potential of single-isomer sulfated beta-cyclodextrin synthesis intermediates in non-aqueous capillary electrophoresis

The enantioselectivity of neutral single-isomer synthetic precursors of sulfated-β-cyclodextrins was studied. Four neutral single-isomer cyclodextrins substituted on the secondary side with acetyl and/or methyl functional groups, heptakis(2-O-methyl-3,6-dihydroxy)-β-cyclodextrin (HM-β-CD), heptakis(2,3-di-O-acetyl-6-hydroxy)-β-cyclodextrin (HDA-β-CD), heptakis(2,3-di-O-methyl-6-hydroxy)-β-cyclodextrin (HDM-β-CD), heptakis(2-O-methyl-3-O-acetyl-6-hydroxy)-β-cyclodextrin (HMA-β-CD), and their sulfated analogs the negatively charged heptakis(2,3-di-O-methyl-6-sulfato)-β-cyclodextrin (HDMS-β-CD) and heptakis(2,3-di-O-acetyl-6-sulfato)-β-cyclodextrin (HDAS-β-CD) were investigated by non-aqueous capillary electrophoresis in the view of enantiodiscrimination for various drugs and related pharmaceutical compounds. The focus of the present work was on the chiral selectivity studies of the neutral derivatives, which are the synthesis intermediates of the sulfated products. The chiral recognition experiments proved that among the neutral compounds the HMA-β-CD shows remarkable enantioselectivity towards chiral guests in non-aqueous capillary electrophoresis, while HM-β-CD, HDA-β-CD and HDM-β-CD failed to resolve any of the 25 studied racemates under the applied experimental conditions. In order to get deeper insight into the molecular interactions between the studied single-isomer cyclodextrin and chiral fluoroquinolones (ofloxacin, gatifloxacin and lomefloxacin) and β-blockers (propranolol), 1H and ROESY NMR experiments were performed. The 2-O-methylation in combination with the 3-O-acetylation of the host was evidenced to exclusively carry the essential spatial arrangement for chiral recognition.

Development of Electrophoretic Methods for Simultaneous Determination of Enantiomeric Ratio and Composition of Diastereomeric Salt Mixtures

Obtaining enantiomeric pure compounds is—among other techniques—possible in a resolvation experiment via diastereomeric salt formation, excellently exemplified by a modified Pope–Peachy method performed in supercritical carbon dioxide as solvent. The salt precipitation is followed by supercritical fluid extraction (SFE) to separate the diastereomeric salts and the unreacted enantiomers. To evaluate the extraction efficiency, conversion and enantioselectivity achieved, it is essential to determine the enantiomer excess and the residual resolving agent content in extracts and raffinates. Carefully chosen experimental parameters enable the simultaneous determination of certain anions and cations in capillary electrophoresis in a single run, which has not been reported for diastereomeric mixtures so far. In this paper, a partially validated chiral selective cyclodextrin enabled capillary electrophoresis method is presented for the characterization of cis-permethrinic acid samples resolved with (R)-1-phenylethylamine prepared by the SFE-based resolvation technique. To evaluate the efficiency of the resolvation, a cyclodextrin enabled chiral separation method was developed applying permethylated-β-cyclodextrin as chiral selector. The theoretical possibility of the widespread application of the developed method (with minor adjustments) is justified for other selectands and selectors. The developed methods can be thereby applied for the fast and reliable control of resolvation experiments.

Synthesis, analytical characterization and capillary electrophoretic use of the single-isomer heptakis-(6-O-sulfobutyl)-beta-cyclodextrin

This contribution reports the synthesis, characterization and capillary electrophoretic application of heptakis-(6-O-sulfobutyl-ether)-β-cyclodextrin sodium salt, (6-(SB)7-β-CD). The compound was obtained through a five-steps synthesis and it represents the first example of single-isomer sulfobutylated cyclodextrin that carries the negatively charged functions exclusively on its primary side and it is unmodified on the lower rim. The purity of each intermediate was determined by appropriate liquid chromatographic methods, while the isomeric purity of the final product was established by an ad-hoc developed HPLC method based on a CD-Screen-IEC column. The structural identification of 6-(SB)7-β-CD was carried out by 1D, 2D NMR spectroscopy and ESI-MS. The chiral separation ability of 6-(SB)7-β-CD was studied by chiral capillary electrophoresis using the single-isomer host as a background electrolyte additive to separate the enantiomers of a representative set of pharmacologically significant model compounds such as verapamil, dapoxetine, ondansetron, propranolol, atenolol, metoprolol, carvedilol, terbutaline, amlodipine and tadalafil. The enantiomer migration order and the effects of the selector concentration on the enantiorecognition properties were investigated. NMR spectroscopy was applied to deepen and further confirm the host-guest interactions and in the case of the model compound dapoxetine a potential representation for the supramolecular assembly was developed based on the dataset collected by the extensive 2D NMR analysis. This single-isomer chiral selector offers a new alternative to the widely applied randomly sulfobutylated- and sulfated-beta-cylodextrins as well as to the single-isomer sulfated and carboxymethylated derivatives in chiral separations.

Stabilization of nanosized titanium dioxide by cyclodextrin polymers and its photocatalytic effect on the degradation of wastewater pollutants

Advanced oxidation processes (AOPs) are considered highly competitive water treatment technologies for the removal of organic pollutants. Among AOP techniques, photocatalysis has recently been the most widely studied. Our aims were to investigate how the dispersion of nanosized titanium dioxide (nanoTiO2) applied in photodegradation-based procedures can be stabilized with cyclodextrins in order to obtain a new, more efficient photocatalyst for the purification of waters polluted by xenobiotics applying UV irradiation. During our work, on the one hand, we studied the behavior and stability of nanoTiO2 in cyclodextrin solutions. On the other hand, we used various monomer and polymer cyclodextrin derivatives, and assessed the options for nanoTiO2 stabilization in the presence of various salts and tap water on the basis of turbidity tests. The physical stability of nanoTiO2 dispersions is diminished in the presence of the salts found in tap water (and occurring also in surface waters and ground water) and they are precipitated immediately. This colloidal instability can be improved by cyclodextrin derivatives. Based on the results of our studies we have selected carboxymethyl β-cyclodextrin polymer (CMBCD-P) for stabilization of nanoTiO2 dispersions. The photocatalytic degradation of methylene blue and ibuprofen as model organic pollutants in various media (distilled water, NaCl solution and tap water) has been studied using nanoTiO2 as catalyst stabilized by CMBCD-P. CMBCD-P itself showed a catalytic effect on the UV degradation of methylene blue. In addition to enhancing the colloid stability of nanoTiO2 CMBCD-P showed also synergistic effects in catalyzing the photodecomposition process of the dye. On the other hand, ibuprofen as a model pharmaceutical, a pollutant of emerging concern (EP), was protected by CMBCD-P against the photocatalytic degradation showing that inclusion complex formation can result in opposite effects depending on the structure of the host–guest complex.

Characterization of a single-isomer carboxymethyl-beta-cyclodextrin in chiral capillary electrophoresis

In this work, the synthesis, characterization, and chiral capillary electrophoretic study of heptakis‐(2,3‐di‐O‐methyl‐6‐O‐carboxymethyl)‐β‐CD (HDMCM), a single‐isomer carboxymethylated CD, are presented. The pH‐dependent and selector concentration‐dependent enantiorecognition properties of HDMCM were investigated and discussed herein. The enantioseparation was assessed applying a structurally diverse set of noncharged, basic, and zwitterionic racemates. The increase in the selector concentration and gross negative charge of HDMCM improved the enantioseparation that could be observed in the majority of the cases. HDMCM was also successfully applied as BGE additive in NACE using a methanol‐based system in order to prove the separation selectivity features and to highlight the broad applicability of HDMCM. Over 25 racemates showed partial or baseline separation with HDMCM under the conditions investigated, among which optimal enantiomer migration order was found for the four stereoisomers of tadalafil, tapentadol, and dapoxetine, offering the possibility of a chiral CE method development for chiral purity profiling of these drugs.