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.     

Retention and selectivity of aromatic hydrocarbons with mono-substituted polar groups in ternary RP-HPLC systems

Summary The variation in selectivity for aromatic hydrocarbons with mono-substituted polar groups is investigated in ternary mobile-phase systems on C₁₈ stationary phases. The dependence of log k′ on the proportions of two modifiers was determined for the solutes within the concentration range of ternary systems obtained by mixing two binary eluents: methanol + water and tetrahydrofuran + water or acetonitrile + water and tetrahydrofuran + water. The nature of the relationships is explained in terms of molecular shape, molecular interactions between solute and extracted modifier and the ordering of solvated ligands on the stationary phase.     

Effect of the alkyl chain length of the bonded stationary phase on solute retention in reversed-phase high-performance liquid chromatography

Summary A comparison of the chromatographic retention characteristics of nonpolar bonded stationary phases was investigated. The results show that the interactions between the solutes and the stationary phases having C₂, C₈ and C₁₈ alkyl groups are almost similar in the range of the mobile phase investigated. This interaction is considered as the solvophobic effect.     

Preparation and evaluation of n-octadecylphosphonic acid-modified magnesia-zirconia stationary phases for reversed-phase liquid chromatography

Three n-octadecylphosphonic acid-modified magnesia-zirconia reversed stationary phases (C₁₈PZM) are prepared via the strong Lewis base interactions between organophosphonate and magnesia-zirconia composite. And two of them are end-capped by using trimethylchlorosilane as end-capping agent in different procedures. Stability studies at extreme high pH conditions (pH 9-12) show that both the non-endcapped and endcapped columns are quite stable at pH 12 mobile phase. The reversed-phase liquid chromatographic behavior of three C₁₈PZM stationary phases are comparatively investigated in detail using a variety of basic compounds as probes. The retention of basic compounds on the three phases is studied over a wide range of pHs. And the possible retention mechanisms of basic compounds on the three stationary phases are discussed. The results show that the basic solutes retain by a hydrophobic and cation-exchange interaction mixed mechanism on three stationary phases when they are operated in eluents at pH values near to the pK_a of the Brönsted conjugate acid form of the analyte, suggesting that inherent zirconol groups on ZM are not expected to interact with bases via cation-exchange interaction at lower pH. Nonetheless, the non-endcapped phase differs markedly from the edncapped ones in retention and selectivity of basic solutes using eluents at pH 4.1, implying a complex retention mechanism at this pH. The cation-exchange sites under such conditions are more likely due to the adsorbed Lewis base anionic buffer constituents (acetate) on accessible ZM surface sites than the chemisorbed phosphonate. Although the three phases exhibit very similar chromatographic behavior with eluents at pH 10.1, and show in general satisfactory separation of basic compounds and alkaloids studied, the performance for a specific analyte, however, differs largely from column to column.     

Comparison of the Separation Characteristics of the Organic–Inorganic Hybrid Octadecyl Stationary Phases XTerra MS C18 and XBridge C18 and Shield RP18 in RPLC

Differences in the system constants of the solvation parameter model, discontinuities in retention factor plots (log k against volume fraction of organic solvent) and retention factor correlation plots are used to study the retention mechanism on XTerra MS C₁₈, XBridge C₁₈ and XBridge Shield RP₁₈ stationary phases with acetonitrile–water and methanol–water mobile phases containing from 10 to 70% (v/v) organic solvent. Wetting of XBridge C₁₈ at 10 and 20% (v/v) acetonitrile is incomplete and is responsible for small changes in the retention mechanism. The intermolecular interactions responsible for retention on XTerra MS C₁₈ and XBridge C₁₈ are similar with minor differences in cavity formation and hydrogen-bonding interactions responsible for small selectivity differences. On the other hand, for bulky solutes there are large changes in retention at low volume fractions of organic solvent (<40% v/v) associated with steric repulsion on the XTerra MS C₁₈ stationary phases that is absent for XBridge C₁₈. Selectivity differences are more apparent for XBridge C₁₈ and XBridge Shield RP₁₈. For acetonitrile-water mobile phases cavity formation in the solvated XBridge Shield RP₁₈ is slightly more difficult and hydrogen-bond acid and base interactions are more important than for XBridge C₁₈. With methanol–water mobile phases it becomes slightly easier to form a cavity in the solvated XBridge RP₁₈ compared with XBridge C₁₈. In addition, the methanol-water solvated XBridge RP₁₈ is a stronger hydrogen-bond base and more dipolar/polarizable than XBridge C₁₈. These variations in selectivity justify the use of XBridge C₁₈ and XBridge Shield RP₁₈ as complementary stationary phases for method development.     

Comparative evaluation of retention of hydrocarbons present in the C5-petroleum fraction on methylsilicone and squalane phases

Summary The retention of hydrocarbons present in the C₅ pyrolysis fraction of gasolines on the stationary phases squalane and methylsilicone oil JXR at 30, 40 and 50°C was investigated by capillary gas chromatography. The unified retention indices of the hydrocarbons were also calculated on squalane. The retention indices obtained on these two phases were interrelated and the quantitative relationship with the structure of the solutes was studied. Equations based on the unified retention indices calculated on squalane and some selected structural elements of the solutes permit the calculation of their retention on methylsilicone with sufficient accuracy.     

Retention of cyanoalkanes and cyanoalkylbenzenes by bonded stationary phases in supercritical fluid chromatography

Summary The retention of cyanoalkanes and cyano-alkylbenzenes during SFC was investigated on alkyl and cyanoalkyl-bonded stationary phases and compared to alkane retention. The particular behaviour of short chain homologues was attributed to a silanophilic interaction with surface-OH groups on the silica and inhibition of specific interactions between solutes and C₁₀CN bonded groups.     

Separation of Dietary Folates by Gradient Reversed-Phase HPLC: Comparison of Alternative and Conventional Silica-Based Stationary Phases

A study of ten silica-based stationary phases and gradient elution conditions to separate dietary folates by reversed-phase HPLC was performed. Alkyl-bonded stationary phases (both conventional and alternative) were found to be the most promising for the separation of different folate monoglutamates in terms of selectivity and peak shape. These phases were better than phenyl-bonded phases which lacked selectivity when separating 10-formyl-folic acid and 5-formyl-tetrahydrofolate. Polar-bonded (cyano) stationary phase showed similar retention characteristics as the conventional alkyl-bonded phases, but ranked below those in terms of peak shape. Overall, alternative stationary phases exhibited slightly higher retention of late-eluted folates and greater retention variability for early-eluting tetrahydrofolate and 5-methyl-tetrahydrofolate. Best selectivity was achieved on alternative polar endcapped Aquasil C₁₈ followed by conventional Synergy MAX C₁₂ and Genesis C₁₈ stationary phases.     

New Fluorous Reversed Phase Silica Gels for HPLC Separations of Perfluorinated Compounds

Two new perfluoroalkyl-modified stationary phases were prepared and compared with two commercially available perfluorinated stationary phases (Fluophase RP and Fluophase Pentafluorophenyl) and a C₁₈-RP column with respect to retention times of an array of perfluoroalkyl-tagged and untagged molecules. A few highly lipophilic compounds were also included in the study. They showed high retention times on C₁₈-RP columns, but not on perfluorinated support materials. Perfluoroalkyl-tagged compounds revealed a weak interaction with the pentafluorophenyl-modified support. The interaction between perfluoroalkyl-tagged compounds and perfluoroalkyl-modified stationary phases was strong, and dependent on the chain length of the perfluoro tags. Surprisingly, there was only a small difference between the retention times of perfluorinated compounds on C₁₈-RP and C₆F₁₃-modified support. Fluorous-fluorous interactions became prevalent only with C₈F₁₇-tagged compounds on C₈F₁₇ functionalized silica gel. Compounds with two perfluoro tags showed a drastic increase in retention time, which might be due to a cooperative effect. These results demonstrate the uniqueness of fluorous-fluorous interactions based on linear perfluoroalkyl chains and open up possibilities for the design of new perfluoro tags for purifications and noncovalent attachments of catalysts or biomolecules on perfluorinated solid supports.     

Correlation between retention and the C18 silica surface coverage in reversed-phase liquid chromatography

Summary Members of three homologous series and five non-homologous solutes with various functional groups were chromatographed on a series of well-characterized C₁₈ reversed stationary phases with a range of methanol-water mobile phases. Measured capacity factors of solutes were correlated with the concentration of C₁₈ ligands on the stationary phase. A linear relationship with the slope depending on a solute molecular structure and the volume fraction of methanol in the mobile phase was obtained. A method for the evaluation of phase ratio is also proposed. Presented at the 17th International Symposium on Chromatography, September 25–30, 1988, Vienna, Austria.     

Synthesis and characterization of novel polar-embedded silica stationary phases for use in reversed-phase high-performance liquid chromatography

We have developed a series of new C₁₀ dipeptide stationary phases via a simple and effective synthetic method. The preparation of the new phases involves the synthesis of silanes and the surface modification of silica. Chromatographic evaluations of these columns were performed using the Engelhardt, Tanaka, and Neue test mixtures. The applicability of these new stationary phases was also evaluated using a series of diagnostic probes including acids, bases or neutral compounds and several generic applications. These new C₁₀ dipeptide stationary phases showed excellent hydrolytic stability over a wide pH range. Like other existing amide-embedded columns, these new stationary phases exhibit higher retention for polar and hydrophilic compounds and different selectivity as compared to conventional C₁₈ columns. These new phases are compatible with 100% aqueous mobile phases, and also provide high column efficiency and good peak shapes for both acidic and basic compounds.     

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.     

Partition and adsorption in the separation of long chain lipids by normal phase high performance liquid chromatography

Martin's adaption of Raoult's Law has been shown to describe partition phenomena in normal phase HPLC. Thus, in the separation of fatty acid pentaflorobenzyl esters and of triacylglycerols in dichloromethane or acetonitrile in half-water-saturated hexane, the natural logarithm of the corrected retention volume is proportional to the number of double bonds that these solutes contain. By extending Martin's adaption of Raoult's Law with activity coefficients, evidence was obtained that deviations observed for the homologous series of saturated fatty acid pentafluorobenzyl esters from C₂ to C₅ could be accounted for by a diminution of adsorption by the inductive effect. For chain lengths longer than C₅, it appeared that there were additional field and inductive effects from the methyl end of the hydrocarbon chains. Thus, with these half-water-saturated solvent systems, the silica surface is probably covered with a bilayer of water which, in the dynamic situation in an HPLC column, attracts several layers of the polar organic component of the mobile phase by dispersive interactions. This stationary phase surface may, then, be considered to be immiscible with the predominantly hexane mobile phase due to these dispersive interactions and thus partition can occur between these two phases. Solutes that enter the stationary phase are then free to interact with the underlying aqueous phase by adsorption.     

Comparison of retention of aromatic hydrocarbons with polar groups in binary reversed-phase high-performance liquid chromatography systems

The retention of aromatic hydrocarbons with polar groups has been correlated as log k1 versus log k2 for reversed-phase high-performance liquid chromatography systems with different binary aqueous mobile phases containing methanol, acetonitrile or tetrahydrofuran as modifiers. Distinct changes in separation selectivity have been observed between tetrahydrofuran and acetonitrile or methanol systems. Methanol and acetonitrile systems show lower diversity of separation selectivity. The changes in retention and selectivity of aromatic hydrocarbons with various polar groups between any two chromatographic systems with binary aqueous eluents (tetrahydrofuran vs. acetonitrile, tetrahydrofuran vs. methanol and methanol vs. acetonitrile) have been interpreted in terms of molecular interactions of the solute with especially one component of the stationary phase region, i.e. extracted modifier, and stationary phase ordering. The ordering of the stationary phase region caused by modifier type influences the chromatographic selectivity of solutes with different molecular shape.     

Characterization of stationary/mobile phase combinations by using markers : Part 1. Preliminary results for the prediction of reversed-phase retention on various stationary phases

The characterization of stationary/mobile phase combinations can be done in a phenomenological way by measuring the k′ values of specific solutes, the markers. These markers can be chosen optimally from a set of test solutes with the use of multivariate techniques. When retention data of solutes on different stationary phases, with varying mobile-phase compositions, are available, a procedure is given to predict the retention of those solutes on new stationary phases. This procedure uses markers to characterize the new stationary/mobile phase combinations.     

Ion‐exchange vs reversed‐phase chromatography for separation and determination of basic psychotropic drugs

Ion exchange chromatography, an alternative to reversed‐phase (RP) chromatography, is described in this paper. We aimed to obtain optimal conditions for the separation of basic drugs because silica‐based RP stationary phases show silanol effect and make the analysis of basic analytes hardly possible. The retention, separation selectivity, symmetry of peaks and system efficiency were examined in different eluent systems containing different types of buffers at acidic pH and with the addition of organic modifiers: methanol and acetonitrile. The obtained results reveal a large influence of the salt cation used for buffer preparation and the type of organic modifier on the retention behavior of the analytes. These results were also compared with those obtained on an XBridge C₁₈ column. The obtained results demonstrated that SCX stationary phases can be successfully used as alternatives to C₁₈ stationary phases in the separation of basic compounds. The most selective and efficient chromatographic systems were applied for the quantification of some psychotropic drugs in fortified human serum samples. Copyright © 2015 John Wiley & Sons, Ltd.     

Retention Behavior of Neutral and Positively and Negatively Charged Solutes on an Immobilized‐Artificial‐Membrane (IAM) Stationary Phase

The retention behavior of neutral, positively charged, and negatively charged solutes on the IAM.PC.DD2 stationary phase was investigated and compared. A set of monofunctional compounds and complex drugs (steroids, nonsteroidal anti‐inflammatory drugs, and β‐blockers) were selected for this study, i.e., neutral solutes and solutes with acidic or basic functionalities which are positively charged or negatively charged at pH 7.0. The correlation between the retention factor log k_w at pH 7.0 on the IAM.PC.DD2 stationary phase and the partition coefficient log P_oct or the distribution coefficient log D_7.0 showed that the retention mechanism depends on the charge state and structural characteristics of the compounds. The neutrals were least retained on the IAM.PC.DD2 stationary phase, and positively charged solutes were more retained than negatively charged ones. This implies that the retention of the charged solutes is controlled not only by lipophilicity but also by the electrostatic interaction with the phospholipid, with which positively charged solutes interact more strongly than negatively charged ones.     

Chromatographic Behaviour and Lipophilicity of Estradiol Derivatives

The retention behaviour of estradiol derivatives has been studied by HPLC on polar chemically bonded stationary phases: C₃CN, DIOL and C₃NH₂, commercially available columns. The mobile phases used were: methanol-water and acetonitrile-water in various proportions. Reversed-phase chromatography occurred on polar chemically bonded stationary phases. Correlation between the retention constants of estradiol derivatives obtained on polar chemically bonded phases and log P calculated via different methods was examined too.     

Further investigations of the effect of pressure on retention in ultra-high-pressure liquid chromatography

In this study, we investigated further the large increases in retention with pressure that we observed previously in RP-LC especially for ionised solutes. These findings were initially confirmed on a conventional silica C₁₈ column, which gave extremely similar results to the hybrid C₁₈ phase originally used. Large increases in retention factor of ∼50% for a pressure increase of 500 bar were also shown for high MW polar but neutral solutes. However, experiments with the same bases in ionised and non-ionised forms suggest that somewhat greater pressure-induced retention increases are found for ionised solutes. Retention increases with pressure were found to be considerably smaller for a C₁ column compared with a C₁₈ column; decreases in retention with increasing pressure were noted for ionised bases when using a bare silica column in the hydrophilic interaction chromatography (HILIC) mode. These observations are consistent with the partial loss of the solvation layer in RP-LC as the solute is forced into the hydrophobic environment of the stationary phase, and consequent reduction in the solute molar volume, while the water layer on the surface of a HILIC packing increases the hydration of a basic analyte. Finally, retention changes with pressure in RP-LC can also be observed at a mobile phase pH close to the solute pK_a, due to changes in pK_a with pressure. However, this effect has no influence on the results of most of our studies.     

An insight into the use of dimethylphenyl carbamate cyclofructan 7 chiral stationary phase in supercritical fluid chromatography: The basic comparison with HPLC

Cyclofructan‐based chiral stationary phases were previously shown as a promising possibility for separation of chiral compounds in high performance liquid chromatography. In this work retention and enantiodiscrimination properties of the 3,5‐dimethylphenyl carbamate cyclofructan 7 chiral stationary phase are described in supercritical fluid chromatography. The results obtained in both of the separation methods were compared. A set of compounds with axial or central chirality was used as analytes. The effect of mobile phase composition, that is, addition of different alcohol modifiers and/or trifluoroacetic acid to carbon dioxide, was examined in the supercritical system. Similarly, mobile phases composed of hexane modified with propan‐2‐ol and/or trifluoracetic acid were used in liquid chromatography. A linear free energy relationship model was utilized for characterization of interactions that are decisive for retention and separation in both techniques. Dispersion interactions showed similar negative values using both methods. The main contribution of hydrogen bond acidity was also comparable for both methods. The propensity to interact with n‐ and/or π‐electron pairs of solutes was significant only in the supercritical system.