Eva Tesařová

Methacrylate monolithic columns of 320 μm I.D. for capillary liquid chromatography

Monolithic capillary columns (320 microm I.D.) were prepared for capillary liquid chromatography (CLC) by radical polymerization of butylmethacrylate (BMA) and ethylenedimethacrylate (EDMA) in the presence of a porogen solvent containing propan-1-ol, butane-1,4-diol and water. The influence of the contents of the porogen solvent and EDMA in the polymerization mixture on the monolith porosity and column efficiency was investigated. The composition of the polymerization mixture was optimized to attain a minimum HETP of the order of tens of microm for test compounds with various polarities. The separation performance and selectivity of the most efficient monolithic column prepared was characterized by van Deemter curves, peak asymmetry factors and Walters hydrophobicity and silanol indices. It was demonstrated that the 320-microm I.D. monolithic column exhibited CLC separation performance similar to that observed for 100- and 150-microm I.D. monolithic columns reported in the literature; moreover, the 320-microm I.D. column was easier to operate in CLC and exhibited a higher sample loadability.

Interactions of basic compounds in reversed-phase high-performance liquid chromatography influence of sorbent character, mobile phase composition, and pH on retention of basic compounds

Differences in a degree of base deactivation of selected reversed-phase sorbents were evaluated. Five silica-based sorbents (SGX C18, LiChrosorb RP-18, LiChrospher RP-select B, Purospher RP-18, Symmetry C18), a polybutadiene-coated alumina (Aluspher RP-select B) and 2-hydroxyethylmethacrylate-based stationary phase (HEMA-BIO 1000 C18) were compared. The best results were obtained for Symmetry C18 and HEMA-BIO 1000 C18. For testing purposes a set of nineteen basic compounds differing in pKa constants and hydrophobicity was used. The bases possessing high pKa values (pKa>9.0) and/or low hydrophobicity proved to be the best indicators of sorbent surface deactivation. Dependencies of capacity factor on pH of the mobile phase were measured. Shape deviations between the sigmoidal theoretically predicted and experimentally obtained curves evidence for a complex retention mechanism in which not only hydrophobic but also other interactions, for example, ion-exchange, participate. The effect of methanol content in the mobile phase on the dissociation of buffer and a basic analyte used was studied. The characteristic shifts of k versus pH of the mobile phase dependencies caused by the addition of methanol were found to be in a good agreement with theory.

Supercritical fluid chromatography as a tool for enantioselective separation; A review

Supercritical fluid chromatography (SFC) has become popular in the field of enantioselective separations. Many works have been reported during the last years. This review covers the period from 2000 till August 2013. The article is divided into three main chapters. The first one comprises a basic introduction to SFC. The authors provide a brief explanation of general principles and possibilities of this method. The advantages and drawbacks are also listed. Next part deals with chiral separation systems available in SFC, namely with the commonly used chiral stationary phases. Properties and interaction possibilities of the chiral separation systems are described. Recent theoretical papers are emphasized in this chapter. The last part of the paper gives an overview of applications of enantioselective SFC in analytical chemistry, in both analytical and preparative scales. Separation systems and conditions are summed up in tables so that they provide a helpful tool for analysts who search for a particular method of analysis.

Gas and high-performance liquid chromatography of phenols

SummaryGas and high-performance liquid chromatographic methods in the analysis of phenols are reviewed. Among the great number of phenolic compounds analyzed, alkylphenols, chlorophenols, dihydroxy- and trihydroxybenzenes and biphenols are chiefly considered. The advantages and drawbacks of the methods are discussed, quoting the most important work. Relationships between the structural characteristics of phenols, the stationary phase structure, the mobile phase composition and the retention data are treated. Typical examples of the conditions for GC and HPLC analysis are summarized in tables. The literature comprises over 270 references, most of them recent.

Comparison of enantioselective separation of N-tert.-butyloxycarbonyl amino acids and their non-blocked analogues on teicoplanin-based chiral stationary phase

Abstract A teicoplanin-based chiral stationary phase (CSP) was tested for enantioseparation of underivatized amino acids and their N - tert .-butyloxycarbonyl ( t -Boc) derivatives, important precursors in peptide synthesis. Mobile phase composition was optimized for organic modifier and triethylamine acetate buffer (TEAA) contents, and retention and enantioresolution of t -Boc-amino acids and their non-blocked analogues were compared. The importance of the amino group of amino acids in the interaction mechanism was evaluated. Native amino acids have better possibility for interaction with teicoplanin; they are more retained on the CSP and better enantioresolved than the blocked amino acids. Presence of triethylamine in the aqueous portion of the mobile phase was shown to be important for separation of enantiomers of t -Boc-amino acids; while native amino acids were almost not affected by the addition of TEAA. Results obtained for blocked amino acid on teicoplanin CSP with mobile phase composed of 1% triethylamine acetate, pH 4.1 and organic modifier were compared to those got on hydroxypropyl-β-cyclodextrin CSP. Good enantioseparation in a reasonable analysis time was obtained on the teicoplanin-based chiral stationary phase.

Recentchiral selectors for separation in HPLC and CE

AbstractEnantiomers (stereoisomers) can exhibit substantially different properties if present in chiral environments. Since chirality is a basic property of nature, the different behaviors of the individual enantiomers must be carefully studied and properly treated. Therefore, enantioselective separations are a very important part of separation science.To achieve the separation of enantiomers, an enantioselective environment must be created by the addition of a chiral selector to the separation system. Many chiral selectors have been designed and used in various fields, such as the analyses of drugs, food constituents and agrochemicals. The most popular have become the chiral selectors and/or chiral stationary phases that are of general use, i.e., are applicable in various separation systems and allow for chiral separation of structurally different compounds.This review covers the most important chiral selectors / chiral stationary phases described and applied in high performance liquid chromatography and capillary electrophoresis during the period of the last three years (2008–2011).

Monolithic columns based on a poly(styrene-divinylbenzene-methacrylic acid) copolymer for capillary liquid chromatography of small organic molecules.

A very simple and readily performed method is described for the preparation of poly(styrene-divinylbenzene-methacrylic acid) monolithic columns for capillary liquid chromatography. The effect of the methacrylic acid content on the morphological and chromatographic properties has been investigated. Methacrylic acid is shown to be essential for isocratic separations of small organic analytes by capillary liquid chromatography. Column efficiencies of about 28,000 theoretical plates/m have been obtained for all the test compounds. The batch-to-batch and run-to-run repeatability of the retention times is better than 1.5%.

Enantioseparation of semisynthetic ergot alkaloids on vancomycin and teicoplanin stationary phases

The macrocyclic antibiotics, vancomycin and teicoplanin, were used as chiral stationary phase selectors for the enantioselective separation of semisynthetic ergot alkaloids in reversed-phase high-performance liquid chromatography (RP-HPLC). The chromatographic behavior of the ergot preparations was investigated in order to obtain a deeper insight into the enantiodiscriminative process. A variety of factors, including mobile phase parameters such as the nature and concentration of the organic modifier, buffer concentration and pH, were examined. Conditions for the enantioseparation of real pharmaceutical preparations, i.e. lisuride, terguride and nicergoline, were found. Differences in the chiral stationary phases are presented and the interaction mechanism is discussed.

Characterization of new R-naphthylethyl cyclofructan 6 chiral stationary phase and its comparison with R-naphthylethyl β-cyclodextrin-based column

Derivatized cyclofructans have been recently introduced as a new class of chiral selectors with great application potential. In this study, a R-naphthylethyl-functionalized cyclofructan 6 based chiral stationary phase (RN CF6 CSP) was used for separation of substituted binaphthyl catalysts in the normal phase HPLC mode. Dominant interaction types that play a role in the separation mechanism were revealed by a linear free energy relationship (LFER) method. In order to evaluate the contribution of the substituent on the cyclofructan structure to retention, the R-naphthylethyl-functionalized β-cyclodextrin (RN CD) CSP was chosen for comparison. Retention factors of 46 widely different solutes, with known solvation parameters, were determined on each of the columns under the same mobile phase compositions used for the enantiomeric separations. The LFER results showed that hydrogen bond acidity and polarity/polarizibility have the greatest impact on retention and enantioresolution on the RN CF6 CSP. The equal influence of the naphthylethyl substituent on the both CSPs was also confirmed while the effects of the basic cyclofructan versus cyclodextrin structures were different. The addition of trifluoroacetic acid to the hexane/propane-2-ol mobile phase was negligible on the RN CF6 CSP for the majority of atropoisomers except for one with ionizable functional groups. The RN CF6 column was shown to be more suitable for enantioseparation of the binaphthyl catalysts than the RN CD column. Higher retention offered by the latter CSP had no positive effect on the enantioresolution.

Enantioselective potential of chiral stationary phases based on immobilized polysaccharides in reversed phase mode.

Derivatized polysaccharide-based columns have high enantiodiscrimination potential mainly in normal phase separation mode. In this work chiral recognition ability of four immobilized polysaccharide-derived chiral stationary phases was evaluated under reversed phase conditions. A set of 30 chiral compounds, particularly drugs possessing various functional groups was used for testing. Baseline enantioseparation was achieved for 17 of them. In general, amylose-based chiral stationary phases showed higher enantioselectivity than the cellulose-based ones, mainly for acidic and bifunctional compounds. The influence of the type and pH of the aqueous mobile phase constituents as well as the role of the organic modifier on the enantioselective separation ability of the stationary phases were also investigated and compared. Complementary separations were obtained on the amylose- and cellulose-based columns. The immobilized polysaccharide-based chiral stationary phases were shown to be useful tool for the enantioseparation of a broad spectrum of chiral analytes in reversed phase separation mode.