Characterization of surface-confined ionic liquid stationary phases: impact of cation and anion identity on retention

A series of surface-confined ionic liquid (SCIL) stationary phases for high-performance liquid chromatography were synthesized in-house. The synthesized phases were characterized by the linear solvation energy relationship (LSER) method to determine the effect of residual linking ligands and the role of the cation and the anion on retention. Statistical analysis was utilized to determine whether the system coefficients returned from multiple linear regression analysis of chromatographic retention data for a set of 28 neutral aromatic probe solutes were significantly different. Examination of the energetics of retention via κ−κ plots agrees with the results obtained from the LSER analysis. Residual linking ligands were determined to contribute reversed-phase-type retention character to the chromatographic system. Furthermore, retention on the SCIL phases was observed to be more profoundly affected by the identity of the anion than by that of the cation.     

Characterization of a novel pyridinium bromide surface confined ionic liquid stationary phase for high-performance liquid chromatography under normal phase conditions via linear solvation energy relationships

Utilizing linear solvation free energy relationship methodology, a novel pyridinium bromide surface confined ionic liquid (SCIL) stationary phase was characterized under normal phase high-performance liquid chromatographic conditions. A limited set of neutral aromatic probe solutes were utilized to rapidly assess the utility of the LSER model, using mobile phases of hexane modified with 2-propanol. The excellent correlation of the global fit across the mobile phase composition range used in this study for the experimental and calculated retention values (R(2)=0.994) indicates that the LSER model is an appropriate model of characterizing this polar bonded phase under normal phase conditions. For a limited subset of compounds, retention on the pyridinium bromide SCIL stationary phase is more highly correlated with that obtained on a cyano column than on a diol column under NP conditions.     

Study of the retention behavior of small polar molecules on different types of stationary phases used in hydrophilic interaction liquid chromatography

The retention behavior of a large group of analytes (35) with varied properties (pK_a and logP) was studied on eight hydrophilic interaction LC columns with different surfaces, stationary phase chemistries, and types of particles. The acetonitrile content (5–95%), buffer concentration (0.5–200 mM), and pH of the mobile phase (3.8 and 6.8) were evaluated for their effects on the retention behavior. The type of stationary phase had a significant impact on the selectivity and retention time of the tested analytes. Completely different selectivity was observed on the aminopropyl stationary phase. In this study, the influence of the buffer concentration was similar for all tested columns, except for the aminopropyl stationary phase. Increasing the buffer concentration led to decreased retention times for the basic compounds and increased retention times for the acidic compounds, while the inverse behavior was observed on the aminopropyl stationary phase. The selectivity of the individual stationary phases was evaluated at pH 3.8 and 6.8. Much lower selectivity differences between the stationary phases were observed at pH 6.8 than pH 3.8. Bare silica stationary phases were used in the comparison of the particles (fused‐core and fully porous particles of 3 and 1.7 μm) and the columns provided by different manufacturers.     

Linear Solvation Energy Relationships in the Determination of Specificity and Selectivity of Stationary Phases

The retention of fifty structurally different compounds has been studied using linear solvation energy relationships. Investigations were performed with the use of six various stationary phases with two mobile phases (50/50 % v/v methanol/water and 50/50 % v/v acetonitrile/water). Packing materials were home-made and functionalized with octadecyl, alkylamide, cholesterol, alkyl-phosphate and phenyl molecules. This is the first attempt to compare all of these stationary phases synthesized on the same silica gel batch. Therefore, all of them may be compared in more complex and believable way, than it was performed earlier in former investigations. The phase properties (based on Abraham model) were used to the classification of stationary phases according to their interaction properties. The hydrophilic system properties s, a, b indicate stronger interactions between solute and mobile phase for most of the columns. Both e and v cause greater retention as a consequence of preferable interactions with stationary phase by electron pairs and cavity formation as well as hydrophobic bonds. However, alkyl-phosphate phase has different retention properties, as it was expressed by positive sign of s coefficient. It may be concluded that most important parameters influencing the retention of compounds are volume and hydrogen bond acceptor basicity. The LSER coefficients showed also the dependency on the type of organic modifier used as a mobile phase component.

     

A Comparison of the Retention Behaviour of Vitamin A and Some Metabolites Eluted from Four Different Reversed Phase High Performance Liquid Chromatographic Stationary Phases

Five retinoids, 13-cis-retinoic acid, 9-cis-retinoic acid, all-trans-retinoic acid, all-trans-retinol and 13-cis-retinal were isocratically separated from four different reversed phase high performance liquid chromatographic stationary phases. By taking advantage of the different retention mechanisms, present between the stationary phases and the analytes, the retinoids were separated with different elution orders using the same mobile phase composition. Two of the stationary phases appeared to have more possibilities to interact with the analytes than the usual hydrophobic interactions. The stationary phase with embedded polar groups showed hydrogen bonding properties and the calix[4]arene based stationary phase showed possibilities to form inclusion complexes with the analytes. These additional interactions appeared to benefit the separations of the analytes. This publication shows the benefits by isocratically separate retinoids employing other stationary phases than the conventional C₁₈ stationary phase.     

Separation of polycyclic aromatic hydrocarbons with a C60 bonded silica phase in microcolumn liquid chromatography

A chemically bonded C₆₀ silica phase was synthesized as a stationary phase for liquid chromatography (LC) and its retention behavior evaluated for various polycyclic aromatic hydrocarbons (PAHs) using microcolumn LC. The results indicate that the C₆₀ bonded phase offers selectivity different from that of octadecylsilica (ODS) bonded phases in the separation of isomeric PAHs. With the C₆₀ phase, PAH molecules having a partial structure similar to that of the C₆₀ molecule, e.g. triphenylene and perylene, were retained longer than with ordinary ODS stationary phases. The results also show that good correlation exists between the retention data with this C₆₀ bonded phase and with C₆₀ itself as the stationary phase.     

Comparison of the Separation Characteristics of the Organic–Inorganic Hybrid Stationary Phases XBridge C8 and Phenyl and XTerra Phenyl in RP-LC

Differences in the system constants of the solvation parameter model and retention factor correlation plots for varied solutes are used to study the retention mechanism on XBridge C₈, XBridge Phenyl and XTerra Phenyl stationary phases with acetonitrile–water and methanol–water mobile phases containing from 10 to 70% (v/v) organic solvent. These stationary phases are compared with XBridge C₁₈ and XBridge Shield RP₁₈ characterized in an earlier report using the same protocol. The XBridge stationary phases are all quite similar in their retention properties with larger difference in absolute retention explained by differences in cohesion and the phase ratio, mainly, and smaller changes in relative retention (selectivity) by the differences in individual system constants and their variation with mobile phase type and composition. None of the XBridge stationary phases are selectivity equivalent but XBridge C₁₈ and XBridge Shield RP₁₈ have similar separation properties, likewise so do XBridge C₈ and XBridge Phenyl, while the differences between the two groups of two stationary phases is greater than the difference within either group. The limited range of changes in selectivity is demonstrated by the high coefficient of determination (>0.98) for plots of the retention factors for varied compounds on the different XBridge phases with the same mobile phase composition.     

Surface-bonded ionic liquid stationary phases in high-performance liquid chromatography—A review

Ionic liquids (ILs) are a class of ionic, nonmolecular solvents which remain in liquid state at temperatures below 100 °C. ILs possess a variety of properties including low to negligible vapor pressure, high thermal stability, miscibility with water or a variety of organic solvents, and variable viscosity. IL-modified silica as novel high-performance liquid chromatography (HPLC) stationary phases have attracted considerable attention for their differential behavior and low free-silanol activity. Indeed, around 21 surface-confined ionic liquids (SCIL) stationary phases have been developed in the last six years. Their chromatographic behavior has been studied, and, despite the presence of a positive charge on the stationary phase, they showed considerable promise for the separation of neutral solutes (not only basic analytes), when operated in reversed phase mode. This aspect points to the potential for truly multimodal stationary phases. This review attempts to summarize the state-of-the-art about SCIL phases including their preparation, chromatographic behavior, and analytical performance.     

Separation behavior of basic compounds on unbonded silicon oxynitride and silica high‐performance liquid chromatography stationary phases with reversed‐phase eluents

Unbonded silicon oxynitride and silica high‐performance liquid chromatography stationary phases have been evaluated and compared for the separation of basic compounds of differing molecular weight, pK_a, and log D using aqueous/organic mobile phases. The influences of percentage of organic modifier, buffer pH, and concentration in the mobile phase on base retention were investigated on unbonded silicon oxynitride and silica phases. The results confirmed that unbonded silicon oxynitride and silica phases demonstrated excellent separation performance for model basic compounds and both the unbonded phases examined possessed a hydrophobic/adsorption and ion‐exchange character. The silicon oxynitride stationary phase exhibited high hydrophilicity compared with silica with a reversed‐phase mobile phase. An ion‐exclusion‐type mechanism becomes predominant for the separation of three aimed bases on the silicon oxynitride column at pH 2.8. Different from silicon oxynitride stationary phase, no obvious change for the retention time of three model bases on silica stationary phase at pH 2.8 can be observed.     

Retention mechanism of the multifunctional solute on columns with different coverage densities using highly aqueous reversed‐phase conditions

A series of 11 homemade octadecyl bonded phases with different coverage densities were tested to determine the influence of the stationary phase on the retention in highly aqueous mobile phases. The concentrations of the organic modifiers (methanol and ACN) were in the range of 0–20%_v/v. The coverage density of bonded ligands and the presence of the end‐capping have strong influence on the solute retention. Amoxicillin (AMO) was chosen as the test compound. Dual properties of AMO, which contain hydrophobic skeleton and polar groups (amino, hydroxyl and carbonyl), cause irregular changes of the retention over the stationary phase hydrophobicity and silanol activity at given mobile phase composition. Presented data show that application of non‐standard low coverage density C18 phases allow to determine AMO in the RPLC condition with high retention.     

Study of RP HPLC Retention Behaviours in Analysis of Carotenoids

For determination of selected carotenoids, various types of columns for high-performance liquid chromatography (HPLC) with different properties have been used. The characteristics of the laboratory-used packing material containing monomeric alkyl-bonded phases (C₁₈, C₃₀) and phenyl as well as phenyl-hexyl stationary phases were studied. The retention data of the examined compounds were used to determine the hydrophobicity and silanol activity of stationary phases applied in the study. The presence of the polar and carboxyl groups in the structure of the bonded ligand strongly influences the polarity of the stationary phase. Columns were compared according to methylene selectivity using a series of benzene homologues. The measurements were done using a methanol–water mobile phase. Knowledge of the properties of the applied stationary phase provided the possibility to predict the RP HPLC retention behaviours in analysis of carotenoids including lutein, lycopene and β-carotene. The composition of the mobile phase, the addition of triethylamine and the type of stationary phase had been taken into account in designing the method of carotenoid identification. Also a monolithic column characterised by low hydrodynamic resistance, high porosity and high permeability was applied. The presented results show that the coverage density of the bonded ligands on silica gel packings and length of the linkage strongly influence the carotenoid retention behaviours. In our study, the highest retention parameters for lutein, lycopene and β-carotene were observed for C₃₀ and C₁₈ stationary phase. This effect corresponds with pore size of column packing greater than 100 Å and carbon content higher than 11 %.     

Click regulation of cyclodextrin primary face for the preparation of novel chiral stationary phases

In this study, a series of novel CD chiral stationary phases were fabricated by immobilization of mono‐6^A‐deoxy‐N₃‐cyclodextrin onto silica surfaces followed by click regulation of CD primary face with 4‐pentynoic acid (acidic moiety), 2‐propynylamine (alkaline moiety) and L‐propargylglycine (chiral amino acid moiety), respectively. Enantioseparations of various kinds of racemates including dansyl‐amino acids, chiral lactides and diketones were conducted in reversed phase modes on these chiral stationary phases, where nearly forty diketones and chiral lactides were firstly separated on cyclodextrin stationary phases. 4‐Pentynoic acid moiety can make the retention ability decline while amine moiety significantly enhanced the retention ability of the stationary phases. For most of the studied analytes, the chiral amino acid moiety had the most positive effects on both the retention time and the resolution. The inclusion complexation between chiral analytes and cyclodextrins were also investigated by fluorescence method.     

Linear Solvation Energy Relationships in the Determination of Specificity and Selectivity of Stationary Phases

The retention of fifty structurally different compounds has been studied using linear solvation energy relationships. Investigations were performed with the use of six various stationary phases with two mobile phases (50/50?% v/v methanol/water and 50/50?% v/v acetonitrile/water). Packing materials were home-made and functionalized with octadecyl, alkylamide, cholesterol, alkyl-phosphate and phenyl molecules. This is the first attempt to compare all of these stationary phases synthesized on the same silica gel batch. Therefore, all of them may be compared in more complex and believable way, than it was performed earlier in former investigations. The phase properties (based on Abraham model) were used to the classification of stationary phases according to their interaction properties. The hydrophilic system properties s, a, b indicate stronger interactions between solute and mobile phase for most of the columns. Both e and v cause greater retention as a consequence of preferable interactions with stationary phase by electron pairs and cavity formation as well as hydrophobic bonds. However, alkyl-phosphate phase has different retention properties, as it was expressed by positive sign of s coefficient. It may be concluded that most important parameters influencing the retention of compounds are volume and hydrogen bond acceptor basicity. The LSER coefficients showed also the dependency on the type of organic modifier used as a mobile phase component.     

Elution order in gas chromatography

The retention behavior of a heterogeneous group of solutes has been examined on seven different stationary phases under isothermal and temperature-programmed conditions. Both ΔH_v (enthalpy of solute vaporization from the stationary phase) and ΔS_v (entropy of solute vaporization from the stationary phase) values were determined for each solute – stationary phase combination under isothermal conditions. Both program rate and carrier gas velocity were shown to affect solute elution order. Unless these and other experimental factors discussed are controlled, column equivalency studies based on solute elution order have dubious value.     

Micellar Liquid Chromatography Study of Quantitative Retention–Activity Relationships for Antihypertensive Drugs

Use of micellar mobile phases in reversed-phase liquid chromatography (RPLC) results in hydrophobic and electrostatic sites for interaction. Modified stationary phases in micellar liquid chromatography (MLC) are structurally similar to biomembranes. To confirm this we focused on the effects of the type and concentration of surfactant (Brij 35, SDS, and CTAB) and mobile phase pH on the retention of antihypertensive drugs on modified C₁₈ stationary phases. Quantitative retention-activity relationships are proposed for the drugs and the different surfactants and compared with those obtained using aqueous–organic mobile phases. Finally, a correlation was obtained between the logarithm of retention factors (log k) and the toxicity (LD₅₀) of antihypertensive drugs. Revised: 14 September 2005 and 4 April 2006     

Chromatography Modeling Software Applicability for Non-Conventional Columns: a Case Study of Calixarene- and Resorcinarene-Bonded Stationary Phases

In this study a systematic evaluation of the applicability of DryLab for calixarene- and resorcinarene-bonded stationary phases and some other relatively new reversed-phase columns with the presence of conventional alkyl-bonded phases was carried out. Calixarene- and resorcinarene-bonded stationary phases belong to the reversed-phase materials. However, depending on the analytes, they show some additional interactions, since their steric, polar and ionic properties are different compared to those of conventional alkyl-bonded phases. Three different mixtures of model analytes, consisting of alkyl substituted benzene derivatives, 4-hydroxybenzoic acid esters and polycyclic aromatic hydrocarbons, were used to verify the accuracy of DryLab prediction of retention times and to compare the results of 20 different liquid chromatographic phases. The type and the content of the organic modifier as well as the temperature and the gradient time were systematically changed using same conditions for all stationary phases. The results showed that the prediction on the calixarene- and resorcinarene-bonded stationary phases as well as on other reversed-phase columns is highly accurate in both isocratic and gradient modes. The predictions and real experiments were highly correlated with an average absolute error (∆t _R) of 0.027 min (<2 s) and an average percent absolute error (%∆t _R) of 0.38 on the calixarene- and resorcinarene-bonded stationary phases, and ∆t _R of 0.04 min (<3 s); %∆t _R of 0.51 on the other reversed-phase columns in this study. As a result, DryLab could be applied with very accurate predictions in method development using calixarene- and resorcinarene-bonded stationary phases, which were used as an example for “new” stationary phase materials.

     

Biparameter Equations for Calculating Kovats Retention Indices of Hydrocarbons

Abstract

Biparameter equations, in which one parameter is the boiling point of the solutes and the other is any of their physicochemical properties able to account for dispersive solute-stationary phase interactions, are obtained by regression analysis. These equations permit the calculation of Kovats retention index (I_R ) for aliphatic and aromatic hydrocarbons in complex mixtures with standard deviations close to experimental error. The linear change of regression coefficients with the work temperature allow us to obtain equations suitable for calculating I_R at any temperature on a given stationary phase. Furthermore, accurate values for the magnitudes included in the equations can be obtained starting from I_R values.

When alkylbenzenes are separated on polar phases, such as Carbowax 20M, it is necessary to add a new parameter accounting for inductive interactions. Once again, regression coefficients, except that of the boiling point which remains constant, change linearly with the polarity of the phase (Tarjan's scale) enabling the obtaining of an equation for calculating I_R on any stationary phase at a given work temperature, although constant deviations between calculated and experimental I_R , on polymeric stationary phases, are found.     

Mobile phase effects on retention on a new butylimidazolium-based high-performance liquid chromatographic stationary phase

A new HPLC stationary phase based on n-butylimidazolium bromide has been characterized by a linear solvation energy relationship (LSER) approach in the binary acetonitrile/water mobile phases. The retention properties of the stationary phase were systematically evaluated in terms of intermolecular interactions between 28 test solutes and the stationary phase. The results and further comparisons with conventional reversed phase system confirm that retention properties are similar to phenyl phases in acetonitrile/water mixtures. The results obtained with acetonitrile/water mixtures are also compared with results obtained using methanol/water mixtures.     

Enantioseparation of chiral sulfonates by liquid chromatography and subcritical fluid chromatography

Tert‐butylcarbamoyl‐quinine and ‐quinidine weak anion‐exchange chiral stationary phases (Chiralpak® QN‐AX and QD‐AX) have been applied for the separation of sodium β‐ketosulfonates, such as sodium chalconesulfonates and derivatives thereof. The influence of type and amount of co‐ and counterions on retention and enantioresolution was investigated using polar organic mobile phases. Both columns exhibited remarkable enantiodiscrimination properties for the investigated test solutes, in which the quinidine‐based column showed better enantioselectivity and slightly stronger retention for all analytes compared to the quinine‐derived chiral stationary phase. With an optimized mobile phase (MeOH, 50 mM HOAc, 25 mM NH₃), 12 of 13 chiral sulfonates could be baseline separated within 8 min using the quinidine‐derivatized column. Furthermore, subcritical fluid chromatography (SubFC) mode with a CO₂‐based mobile phase using a buffered methanolic modifier was compared to HPLC. Generally, SubFC exhibited slightly inferior enantioselectivities and lower elution power but also provided unique baseline resolution for one compound.