Effect of short-chain alcohols on surfactant-mediated reversed-phase liquid chromatographic systems

The behaviour of β-blockers in a reversed-phase liquid chromatographic (RPLC) column with mobile phases containing a short-chain alcohol (methanol, ethanol or 1-propanol), with and without the surfactant sodium dodecyl sulphate (SDS), was explored. Two surfactant-mediated RPLC modes were studied, where the mobile phases contained either micelles or only surfactant monomers at high concentration. Acetonitrile was also considered for comparison purposes. A correlation was found between the effects of the organic solvent on micelle formation (monitored by the drop weight procedure) and on the nature of the chromatographic system (as revealed by the retention, elution strength and peak shape of β-blockers). When SDS is added to the mobile phase, the free surfactant monomers bind the C18 bonded chains on the stationary phase, forming an anionic layer, which attracts strongly the cationic β-blockers. The retention is modified as a consequence of the solving power of the organic solvent, micelles and surfactant monomers. The molecules of organic solvent bind the micelles, modify their shape, and may avoid their formation. They also bind the monomers of surfactant, desorbing them from the stationary phase, which affects the retention. The remaining surfactant covers the free silanols on the siliceous support, avoiding the interaction with the cationic solutes. The retention of β-blockers results from a combination of electrostatic and hydrophobic interactions, the latter being weaker compared to the hydro-organic system. The peak efficiencies and asymmetries are excellent tools to probe the surfactant layer on the stationary phase in an SDS/organic solvent system. The peaks will be nearly symmetrical wherever enough surfactant coats the stationary phase (up to 60% methanol, 40% ethanol, 35% 1-propanol, and 50% acetonitrile).     

Comparison of the performance of non-ionic and anionic surfactants as mobile phase additives in the RPLC analysis of basic drugs

Surfactants added to the mobile phases in reversed-phase liquid chromatography (RPLC) give rise to a modified stationary phase, due to the adsorption of surfactant monomers. Depending on the surfactant nature (ionic or non-ionic), the coated stationary phase can exhibit a positive net charge, or just change its polarity remaining neutral. Also, micelles in the mobile phase introduce new sites for solute interaction. This affects the chromatographic behavior, especially in the case of basic compounds. Two surfactants of different nature, the non-ionic Brij-35 and the anionic sodium dodecyl sulfate (SDS) added to water or aqueous-organic mixtures, are here compared in the separation of basic compounds (β-blockers and tricyclic antidepressants). The reversible/irreversible adsorption of the monomers of both surfactants on the stationary phase was examined. The changes in the nature of the chromatographic system using different columns and chromatographic conditions were followed based on the changes in retention and peak shape. The study revealed that Brij-35 is suitable for analyzing basic compounds of intermediate polarity, using "green chemistry", since the addition of an organic solvent is not needed and Brij-35 is a biodegradable surfactant. In contrast, RPLC with hydro-organic mixtures or mobile phases containing SDS required high concentrations of organic solvents.     

Chromatographic Behavior of Aromatic Diamines in Hydro-Organic, Micellar and Submicellar Reversed Phase Liquid Chromatographic Modes

The chromatographic behavior (retention, elution strength, efficiency, peak asymmetry and selectivity) of some aromatic diamines in the presence of methanol with or without anionic surfactant SDS in the four different reversed phased liquid chromatographic (RPLC) modes, i.e., hydro-organic, micellar (MLC), low submicellar (LSC) and high submicellar (HSC), was investigated. In the three surfactant-mediated modes, the surfactant monomers coat the stationary phase even up to 70 % methanol; this results in the suppression of peak tailing (by masking the silanol groups on the stationary phase). In MLC and HSC, the solute retention decreases by increasing the surfactant concentration, while this situation was reversed in LSC. In the region between MLC and HSC modes (25–50 % methanol), retention of late eluting solutes was increased by increasing methanol content which is seemingly due to disaggregation of SDS micelles. Changes in selectivity were observed after changing the concentrations of SDS and methanol, in a greater extent when concentration of SDS was changed. Among the four studied RPLC modes, HSC showed the best efficiency with nearly symmetrical peaks. Prediction of retention of solutes in HSC based on a mechanistic retention model combined with Pareto-optimality method allowed the full resolution of target diamines in practical analysis times.     

Description of retention characteristics of calixarene-bonded stationary phases in dependence of the methanol content in the mobile phase

Calixarene-bonded stationary phases in HPLC are known to support additional interactions compared to conventional alkyl-bonded phases (π–π interactions, complex-building interactions). Thus it cannot be presumed that the same mechanisms of retention apply and that retention can be predicted in similar ways. Here 31 solutes of highly various molecular structures have been analysed at different mobile phase compositions (0–98% (v/v) methanol) in order to characterise the chromatographic behaviour of the novel stationary phases and to test the applicability of established models predicting retention factors. The influence of a change of the methanol content is discussed for non-polar, polar and ionic solutes and differences of their behaviour on the differing column types are shown. Additionally estimates about underlying retention mechanisms are given.     

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     

Selectivity of calixarene‐bonded silica phases in HPLC: Description of special characteristics with a multiple term linear equation at different methanol concentrations

Retention and selectivity characteristics of different calixarene‐, resorcinarene‐ and alkyl‐bonded stationary phases are examined by analyzing a set of test solutes covering the main interactions (hydrophobic, steric, ionic, polar) that apply in HPLC. Therefore Dolan and Snyder's multiple term linear equation has been adapted to fit the properties of calixarene‐bonded columns. The obtained parameters are used to describe retention and selectivity of the novel Caltrex^® phases and to elucidate underlying mechanisms of retention. Here, differences of stationary phase characteristics at different methanol concentrations in the mobile phases are examined. Both selectivity and retention were found to depend on the methanol content. Differences of these dependencies were found for different stationary phases and interactions. The differences between common alkyl‐bonded and novel calixarene‐bonded phases increase with increasing methanol content.     

阴离子型胶束液相色谱的溶质保留行为

以SDS阴离子表面活性剂作流动相,酸性、中性及两性药物为受试药物,运用三相平衡理论考察影响阴离子型胶束液相色谱（AMLC）溶质保留行为的几个因素。保留由溶质与胶束相及修饰后固定相的综合作用决定。有机调节剂正丙醇的加入改变了溶质从水相到固定相或到胶束相的平衡,保留取决于溶质疏水性和静电性间的平衡。此外对羟基苯甲酸酯类同系物的亲脂性与3种细菌最小抑菌浓度具有显著相关性,提示其抑菌机理主要取决于药物与生物膜的亲和性。     

Characterisation of stationary phases in subcritical fluid chromatography with the solvation parameter model. III. Polar stationary phases

In this third paper, varied types of polar stationary phases, namely silica gel (SI), cyano (CN)- and amino-propyl (NH2)-bonded silica, propanediol-bonded silica (DIOL), poly(ethylene glycol) (PEG) and poly(vinyl alcohol) (PVA), were investigated in subcritical fluid mobile phase. This study was performed to provide a greater knowledge of the properties of these phases in SFC, and to allow a more rapid and efficient choice of polar stationary phase in regard of the chemical nature of the solutes to be separated. The effect of the nature of the stationary phase on interactions between solute and stationary phases and between solute and carbon dioxide-modifier mobile phases was studied by the use of a linear solvation energy relationship (LSER), the solvation parameter model. The retention behaviour observed with sub/supercritical fluid with carbon dioxide-methanol is close to the one reported in normal-phase liquid chromatography with hexane. The hydrogen bond acidity and basicity, and the polarity/polarizability favour the solute retention when the molar volume of the solute reduces it. As with non-polar phases, the absence of water in the subcritical fluid allows the solute/stationary phase interactions to play a greater part in the retention behaviour. As expected, the DIOL phase and the bare silica display a similar behaviour towards acidic and basic solutes, when interactions with basic compounds are lower with the NH2 phase. On the CN phase, all interactions (hydrogen bonding, dipole-dipole and charge transfer) have a nearly equivalent weight on the retention. The polymeric phases, PEG and PVA, provide the most accurate models, possibly due to their better surface homogeneity.     

Submicellar and micellar reversed-phase liquid chromatographic modes applied to the separation of β-blockers

The behaviour of a reversed-phase liquid chromatographic (RPLC) system (i.e. elution order, resolution and analysis time), used in the analysis of β-blockers with acetonitrile–water mobile phases, changes drastically upon addition of an anionic surfactant (sodium dodecyl sulphate, SDS). Surfactant monomers cover the alkyl-bonded phase in different extent depending on the concentration of both modifiers, in the ranges 1 × 10⁻³–0.15 M SDS and 5–50% acetonitrile. Meanwhile, the surfactant is dissolved in the mobile phase as free monomers, associated in small clusters or forming micelles. Four characteristic RPLC modes are yielded, with transition regions between them: hydro-organic, micellar, and low and high submicellar. The mobile phases in the two latter modes contain a concentration of SDS below or well above the critical micellar concentration (CMC) in water (i.e. 8 × 10⁻³ M), and more than 30% acetonitrile. High submicellar RPLC appeared as the most promising mode, as it allowed full resolution of the β-blockers in practical times, while these were unresolved or highly retained in the other RPLC modes. The strong attraction of the cationic solutes to the anionic SDS makes a direct transfer mechanism between surfactant molecules in the stationary and mobile phases likely.     

Insights into the retention mechanism on an octadecylsiloxane-bonded silica stationary phase (HyPURITY C18) in reversed-phase liquid chromatography

Plots of the retention factor against mobile phase composition were used to organize a varied group of solutes into three categories according to their retention mechanism on an octadecylsiloxane-bonded silica stationary phase HyPURITY C18 with methanol-water and acetonitrile-water mobile phase compositions containing 10-70% (v/v) organic solvent. The solutes in category 1 could be fit to a general retention model, Eq. (2), and exhibited normal retention behavior for the full composition range. The solutes in category 2 exhibited normal retention behavior at high organic solvent composition with a discontinuity at low organic solvent compositions. The solutes in category 3 exhibited a pronounced step or plateau in the middle region of the retention plots with a retention mechanism similar to category 1 solutes at mobile phase compositions after the discontinuity and a different retention mechanism before the discontinuity. Selecting solutes and appropriate composition ranges from the three categories where a single retention mechanism was operative allowed modeling of the experimental retention factors using the solvation parameter model. These models were then used to predict retention factors for solutes not included in the models. The overwhelming number of residual values [log k (experimental) - log k (model predicted)] were negative and could be explained by contributions from steric repulsion, defined as the inability of the solute to insert itself fully into the stationary phase because of its bulkiness (i.e., volume and/or shape). Steric repulsion is shown to strongly depend on the mobile phase composition and was more significant for mobile phases with a low volume fraction of organic solvent in general and for mobile phases containing methanol rather than acetonitrile. For mobile phases containing less than about 20 % (v/v) organic solvent the mobile phase was unable to completely wet the stationary phase resulting in a significant change in the phase ratio and for acetonitrile (but less so methanol) changes in the solvation environment indicated by a discontinuity in the system maps.     

Adjusting the chromatographic properties of poly(ionic liquid)‐modified stationary phases by substitution on the imidazolium cation

Poly(ionic liquid)‐modified stationary phases can have multiple interactions with solutes. However, in most stationary phases, separation selectivity is adjusted by changing the poly(ionic liquid) anions. In this work, two poly(ionic liquid)‐modified silica stationary phases were prepared by introducing the cyano or tetrazolyl group on the pendant imidazolium cation on the polymer chains. Various analytes were selected to investigate their mechanism of retention in the stationary phases using different mobile phases. Two poly(ionic liquid)‐modified stationary phases can provide various interactions toward solutes. Compared to the cyano‐functionalized poly(ionic liquid) stationary phase, the tetrazolyl‐functionalized poly(ionic liquid) stationary phase provides additional cation‐exchange and π‐π interactions, resulting in different separation selectivity toward analytes. Finally, applicability of the developed stationary phases was demonstrated by the efficient separation of nonsteroidal anti‐inflammatory drugs.     

Retention characteristics of a new butylimidazolium-based stationary phase. Part II: anion exchange and partitioning

A surface-confined ionic liquid (SCIL) and a commercial quaternary amine silica-based stationary phase were characterized employing the linear solvation energy relationship (LSER) method in binary methanol/water mobile phases. The retention properties of the stationary phases were evaluated in terms of intermolecular interactions between 28 test solutes and the stationary phases. The comparison reveals a difference in the hydrophobic and hydrogen bond acceptance interaction properties between the two phases. The anion exchange retention mechanism of the SCIL phase was demonstrated using nucleotides. The utility of the SCIL phase in predicting logk _IL/water values by chromatographic methods is also discussed.     

Characterisation of stationary phases in subcritical fluid chromatography with the solvation parameter model IV. Aromatic stationary phases

The purpose of the present work was to systematically study the chromatographic behaviour of different aromatic stationary phases in a subcritical fluid mobile phase. We attempted to assess the chemical origin of the differences in retention characteristics between the different columns. Various types of aromatic stationary phases, all commercially available, were investigated. The effect of the nature of the aromatic bonding on interactions between solute and stationary phases and between solute and carbon dioxide-methanol mobile phase was studied by the use of a linear solvation energy relationship (LSER): the solvation parameter model. This study was performed to provide a greater knowledge of the properties of these phases in subcritical fluid chromatography, and to allow a more rapid and efficient choice of aromatic stationary phase in regard of the chemical nature of the solutes to be separated. Charge transfer interactions naturally contribute to the retention on all these stationary phases but are completed by various other types of interactions, depending on the nature of the aromatic group. The solvation vectors were used to compare the different phase properties. In particular, the similarities in the chromatographic behaviour of porous graphitic carbon (PGC), polystyrene-divinylbenzene (PS-DVB) and aromatic-bonded silica stationary phases are evidenced.     

线性参数logI对反相液相色谱中的非极性小分子溶质体系的表征

从理论上阐明了计量置换平衡常数的对数ｌｏｇＫａ对液相色谱中计量置换保留模型的线性参数ｌｏｇＩ起着主导作用。ｌｏｇＩ表示溶质对固定相的亲合势，且具有热力学平衡常数的性质。     

New Perspectives in Micellar Liquid Chromatography

Abstract

This paper will summarize several new findings obtained in our laboratory on the use of micellar mobile phases in liquid chromatography. The topics to be addressed include (i) stationary phase modification by the mobile phase surfactant in micellar liquid chromatography, (ii) investigation of the retention mechanism in micellar liquid chromatography (MLC) using an alkyl-benzene homologous series, (iii) evaluation of the effects of organic additives upon retention and efficiency in MLC, and (iv) preliminary characterization of several new classes of surfactant molecules for use in MLC. The information gained from these studies provides new insights into the dynamics of MLC and demonstrates their potential usefulness in several new separation applications including the resolution of optical isomers.     

Inorganic salts as hold-up time markers in C(18) columns

A systematic study of the retention behaviour in a C(18) reversed phase liquid chromatography (RPLC) system of several inorganic salts, namely NaNO(2), NaNO(3), KBr, Bu(4)NBr and K(2)Cr(2)O(7), is presented. The behaviour of the markers in unbuffered mobile phases and the marker retention dependence on the pH and ionic strength of the buffered mobile phases have been analyzed. In addition, a comparison between the retention behaviour of the markers and several ionizable analytes in buffered eluents is presented.     

Selectivity of some basic solutes on a poly(methyltetradecylsiloxane)-silica stationary phase

Complex analyses of polar compounds, especially basic ones, require more selective stationary phases. The present paper describes a stationary phase prepared by thermal immobilization of poly(methyltetradecylsiloxane) onto chromatographic silica (PMTDS-SiO(2)). This stationary phase presents hydrophobic and ion-exchange interactions that confer both high retention and unique selectivities for basic solutes. The influence of ion-exchange interactions is confirmed by the increase in retention factors of basic solutes when the mobile-phase pH changes from acidic to neutral and by the decrease in retention factors when the mobile-phase pH changes from neutral to alkaline. The ion-exchange properties of the stationary phase are enriched in neutral mobile phase (pH 7-7.5) using soft Lewis bases such as tricine and tris as buffers but are suppressed in both acidic (pH 2.5-6) and highly alkaline mobile phases (pH≤10). Increasing both temperature and flow rate permits more rapid separations while maintaining the selectivity. The stability of the stationary phase is evaluated with acid, neutral and alkaline mobile phases.     

Effect of ionization and the nature of the mobile phase in quantitative structure-retention relationship studies

The octanol-water distribution constant, commonly called partition coefficient, Po/w, is a parameter often retained as a measure of the hydrophobicity of a molecule. log Po/w, for a given molecule, can be conveniently evaluated constructing correlation lines between standard retention factor logarithms (log k) in reversed-phase liquid chromatography (RPLC) and standard log Po/w values. Many compounds of pharmaceutical interest can be quite hydrophobic and have, simultaneously, basic nitrogen atoms or acidic sulfur containing groups in their structure. This renders them ionizable. The hydrophobicity of the molecular drug form (Po/w value) is completely different from its ionic form (log Po/w(+ or -) value). The actual hydrophobicity of such ionizable molecule depends on the pH. It can be represented by an apparent Papp value that takes into account the amount of compound in its molecular and ionic state combining the Po/w and Po/w(+ or -) values. In this work, log k in RPLC for ionizable as well as non-ionizable pharmaceutical compounds with different therapeutic properties (10 beta-blockers, seven tricyclic antidepressants (TA), eight steroids and 12 sulfonamides) were correlated with log Po/w. Similar correlations were done between log k and the corrected log Papp values at pH 3. Aqueous-organic mobile phases containing acetonitrile (conventional RPLC) and micellar-organic mobile phases (micellar liquid chromatography, MLC), prepared with the anionic surfactant sodium dodecyl sulfate and the organic solvents acetonitrile, propanol or pentanol, were also used to elute the compounds. All mobile phases were buffered at pH 3. Using conventional retention RPLC data, the correlation of log k with log Po/w, was satisfactory for steroids because they cannot ionize. For ionizable beta-blockers and TAs, the use of log Papp values improved the quality of the correlations, but yielded similar results for sulfonamides. In MLC, since an electrostatic interaction is added to hydrophobic forces, poorer correlations were obtained in all cases. The retention data obtained in RPLC also seems to correlate better with the biological activity of the drugs.