M.J. Ruiz-Ángel

Retention mechanisms in micellar liquid chromatography

Micellar liquid chromatography (MLC) is a reversed-phase liquid chromatographic (RPLC) mode with mobile phases containing a surfactant (ionic or non-ionic) above its critical micellar concentration (CMC). In these conditions, the stationary phase is modified with an approximately constant amount of surfactant monomers, and the solubilising capability of the mobile phase is altered by the presence of micelles, giving rise to diverse interactions (hydrophobic, ionic and steric) with major implications in retention and selectivity. From its beginnings in 1980, the technique has evolved up to becoming a real alternative in some instances (and a complement in others) to classical RPLC with hydro-organic mixtures, owing to its peculiar features and unique advantages. This review is aimed to describe the retention mechanisms (i.e. solute interactions with both stationary and mobile phases) in an MLC system, revealed in diverse reports where the retention behaviour of solutes of different nature (ionic or neutral exhibiting a wide range of polarities) has been studied in a variety of conditions (with ionic and non-ionic surfactants, added salt and organic solvent, and varying pH). The theory is supported by several mechanistic models that describe satisfactorily the retention behaviour, and allow the measurement of the strength of solute-stationary phase and solute-micelle interactions. Suppression of silanol activity, steric effects in the packing pores, anti-binding behaviour, retention of ionisable compounds, compensating effect on polarity differences among solutes, and the contribution of the solvation parameter model to elucidate the interactions in MLC, are commented.

New Insights and Recent Developments in Micellar Liquid Chromatography

Abstract: Micellar liquid chromatography (MLC) is an efficient alternative to conventional reversed–phase liquid chromatography with hydro‐organic mobile phases. Almost three decades of experience have resulted in an increasing production of analytical applications. Current concern about the environment also reveals MLC as an interesting technique for “green” chemistry because it uses mobile phases containing 90% or more water. These micellar mobile phases have a low toxicity and are not producing hazardous wastes. After a rapid overview of the two first decades of the technique, this review focuses on the recent advances on fundamental aspects and analytical applications. Traditional and new surfactants, search of new organic solvents as mobile phase modifiers, and the use of new columns are addressed. Surfactant‐bonded phase association, combination of diverse surfactant effects, interaction between organic solvents and micelles, and resolution performance are also considered. A special attention has been paid to the limited efficiency and weak elution strength, which are the main limitations usually pointed out in MLC. An effort has been made to clarify some wrong and sometimes unjustified ideas about MLC. The potential of this chromatographic mode is also shown for routine analytical procedures.

Countercurrent chromatography: People and applications

The scientific literature was scanned for the published research articles dealing with countercurrent chromatography (CCC) over the time period 1980-May 2008. The search returned 1638 articles that were analyzed focussing on people and applications. Concerning the people, it was found that the geographical location of the CCC authors was relatively well balanced between USA, Asia with mainly China and Japan and Europe. Yoichiro Ito, the inventor of the technique, is by far the most productive author in the field with 331 articles or more than one over five CCC articles published in the time period. Without surprise, English is the dominant language with more than 82% of the articles. A significant 8% amount of CCC articles were published in Chinese in Chinese journals. Chromatography journals are the logical tribune for half of the published CCC articles. Concerning the applications, the separation and purification of natural compounds is the dominant theme in CCC making the subject of more than one article over two. Starting from the plant extract, CCC in few steps can produce significant amounts of more than 95% pure compounds used for identification and/or property studies. Other applications are found in the pharmaceutical and chemical field. The separation of enantiomers on the preparative scale is a field of growing importance.

Micellar liquid chromatography: suitable technique for screening analysis.

The screening capability of micellar liquid chromatography (MLC) is discussed using the reported chromatographic data of several sets of compounds (amino acids, beta-blockers, diuretics, phenethylamines, phenols, polynuclear aromatic hydrocarbons, steroids and sulfonamides) and new results (sulfonamides and steroids). The chromatographic data are treated with an interpretive optimisation resolution procedure to obtain the best separation conditions. Usually, the pH and the concentration of surfactant (sodium dodecyl sulfate, SDS, or cetyltrimethylammonium bromide) for the optimal mobile phase were 2.5-3 and < 0.12 M, respectively. The nature and concentration of organic solvent depended on the polarity of the eluted compounds: a low volume fraction of propanol (approximately 1%, v/v) was useful to separate the amino acids, with log P(o/w) < -1 (where P(o/w) is the octanol-water partition coefficient). A greater concentration of this solvent (approximately 5-7%) was needed for compounds in the range -1 < log P(o/w) < 2, as with the studied diuretics and sulfonamides, and a high concentration of propanol (approximately 15%) or a low concentration of butanol (< 10%) had to be used for less polar compounds with 1 < log P(o/w) < 3, such as the beta-blockers. Pentanol (< 6%) was more suitable for the even less polar compounds with log P(o/w) > 3, such as the steroids. For basic drugs such as the phenethylamines (0 < log P(o/w) < 1.7), eluted with a micellar eluent of anionic SDS, propanol was too weak. A study is also shown for mixtures of sulfonamides (log P(o/w) = -1.2 to 1.7) and steroids (log P(o/w) = 3.0-8.1) eluted from conventional C18 columns with SDS mobile phases containing acetonitrile and 1-pentanol, respectively, which are compared with classical acetonitrile-water and methanol-water mixtures. The results complement a previous study on beta-blockers (log P(o/w) = -0.03 to 2.8) and reveal that MLC is a very competitive technique for the screening of compounds against conventional RPLC, due to its peculiar behaviour with regard to the selectivity and elution strength. The concentration of organic solvent needed to obtain sufficiently low retention times (even for highly hydrophobic steroids with log P(o/w) = 7-8) is also appreciably smaller for MLC, which reduces the environmental impact of the mobile phases.

On the use of ionic liquids as mobile phase additives in high-performance liquid chromatography. A review

The popularity of ionic liquids (ILs) has grown during the last decades in several analytical separation techniques. Consequently, the number of reports devoted to the applications of ILs is still increasing. This review is focused on the use of ILs (mainly imidazolium-based associated to chloride and tetrafluoroborate) as mobile phase additives in high-performance liquid chromatography (HPLC). In this approach, ILs just function as salts, but keep several kinds of intermolecular interactions, which are useful for chromatographic separations. Both cation and anion can be adsorbed on the stationary phase, creating a bilayer. This gives rise to hydrophobic, electrostatic and other specific interactions with the stationary phase and solutes, which modify the retention behaviour and peak shape. This review updates the advances in this field, with emphasis on topics not always deeply considered in the literature, such as the mechanisms of retention, the estimation of the suppressing potency of silanols, modelling and optimisation of the chromatographic performance, and the comparison with other additives traditionally used to avoid the silanol problem.

Optimised procedures for the reversed-phase liquid chromatographic analysis of formulations containing tricyclic antidepressants

The chromatographic behaviour (retention, selectivity, peak shape and resolution) of seven tricyclic antidepressants (TCAs), amitryptiline, clomipramine, doxepin, imipramine, maprotiline, nortryptiline and trimipramine, was examined. Conventional unendcapped Cs and C18 columns and an endcapped XTerra MS C18 column recommended for the analysis of basic compounds were used together with acetonitrile-water and micellar sodium dodecylsulfate (SDS)-pentanol mobile phases. The two best combinations were XTerra C18/acetonitrile, which yielded the largest efficiencies and resolution, and C8/SDS-pentanol, which eliminated the peak tails that were still observed with the XTerra C18 column. Both the systems were used to develop simple chromatographic procedures for the control of TCAs in pharmaceutical formulations using UV detection. The selected mobile phase compositions were 35% (v/v) acetonitrile (XTerra C18 column) and 0.075 M SDS-6% (v/v) pentanol (C8 column), both at pH 3. Satisfactory recoveries were achieved in both cases, with intra- and inter-day relative standard deviations (RSDs) always below 0.6 and 2.0%, respectively. The preparation of the samples was simple in both modes, since a previous extraction of the drugs was not needed. The micellar mode has, however, the advantage of using a smaller amount of organic solvent, which is retained in the micellar SDS solution. The C8 column is also less expensive.

Retention Mechanisms for Basic Drugs in the Submicellar and Micellar Reversed-Phase Liquid Chromatographic Modes

The reversed-phase liquid chromatographic (RPLC) behavior (retention, elution strength, selectivity, efficiency, and peak asymmetry) for a group of basic drugs (beta-blockers), with mobile phases containing the anionic surfactant sodium dodecyl sulfate (SDS) and acetonitrile, revealed different separation environments, depending on the concentrations of both modifiers: hydro-organic, submicellar at low surfactant concentration and high concentration of organic solvent, micellar, and submicellar at high concentration of both surfactant and organic solvent. In the surfactant-mediated modes, the anionic surfactant layer adsorbed on the stationary phase interacts strongly with the positively charged basic drugs increasing the retention and masks the silanol groups that are the origin of the poor efficiencies and tailing peaks in hydro-organic RPLC with conventional columns. Also, the strong attraction between the cationic solutes and anionic SDS micelles or monomers in the mobile phase enhances the solubility and allows a direct transfer mechanism of the cationic solutes from micelles to the modified stationary phase, which has been extensively described for highly hydrophobic solutes.

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 beta-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 x 10(-3)-0.15M 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 x 10(-3)M), and more than 30% acetonitrile. High submicellar RPLC appeared as the most promising mode, as it allowed full resolution of the beta-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.

Peak half-width plots to study the effect of organic solvents on the peak performance of basic drugs in micellar liquid chromatography

The addition of the anionic surfactant sodium dodecyl sulphate (SDS) to hydro-organic mixtures of methanol, ethanol, propanol or acetonitrile with water yielded enhanced peak shape (i.e. increased efficiencies and symmetrical peaks) for a group of basic drugs (beta-blockers) chromatographed with a Kromasil C18 column. The effect can be explained by the thin layer of surfactant associated to the hydrocarbon chain on the stationary phase in the presence of the organic solvents, which covers the free silanols on the siliceous support avoiding their interaction with the cationic basic drugs. These instead interact with the anionic head of the surfactant increasing their retention and allowing a more facile mass transfer. The peak shape behaviour with the four organic solvents (methanol, ethanol, propanol and acetonitrile) was checked in the presence and absence of SDS. The changes in peak broadening rate and symmetry inside the chromatographic column were assessed through the construction of peak half-width plots (linear relationships between the left and right half-widths at 10% peak height versus the retention time). The examination of the behaviour for a wide range of compositions indicated that the effect of acetonitrile in the presence of SDS is different from ethanol and propanol, which behave similarly. Acetonitrile seems to be superior to the alcohols in terms of peak shape, which can be interpreted by the larger reduction in the adsorbed surfactant layer on the C18 column. However, the decreased efficiencies observed at increasing surfactant concentration in the mobile phase should be explained by the reduction in retention times, more than by a change in the stationary phase nature.

Micellar-organic versus aqueous-organic mobile phases for the screening of β-blockers

A comparative study of the performance of reversed-phase liquid chromatography with micellar-organic (MLC) and aqueous-organic (RPLC) mobile phases is reported for the separation of 16 -blockers (acebutolol, alprenolol, atenolol, bisoprolol, carteolol, celiprolol, esmolol, labetalol, metoprolol, nadolol, oxprenolol, pindolol, practolol, propranolol, sotalol, and timolol). MLC with hybrid mobile phases of sodium dodecyl sulfate (SDS) and propanol is revealed as a very competitive technique for the screening of these drugs. Using a conventional Spherisorb C18 column, the theoretical plates ( N) and asymmetry factors (B/A) for the optimal mobile phase compositions were in the ranges N = 2200–4400 and B/A = 1.0–1.3 for SDS–propanol against N = 290–960 and B/A = 2.2–3.6, and N = 1700–5100 and B/A = 1.5–2.2 for acetonitrile–water in the absence and presence of 0.1% (v/v) triethylamine. With a base-deactivated XTerra MS C18 column and acetonitrile–water, peak shape was improved (N = 1100–8800 and B/A = 1.1–2.5), but the elution pattern (large differences in retention between polar and low polar compounds) was similar to that achieved with the Spherisorb column. A set of 14 -blockers showing a wide range of polarities (log Po/w =− 0.03 for atenolol to 2.60 for propranolol) could be resolved isocratically with a mobile phase of 0.10 M SDS–15% (v/v) propanol with an analysis time of 32 min. The resolution was better than with aqueous-organic RPLC using the conventional and base-deactivated C18 columns, where only 10–12 compounds could be separated in a single chromatogram. Using specific mobile phases of SDS–propanol, all possible binary mixtures of the 16 -blockers could be resolved fully (peak purity of the peak pairs, R> 0.999), except practolol–sotalol (R = 0.992), nadolol–pindolol (R = 0.959), and esmolol–metoprolol (R = 0.956), which showed a small overlapping. The resolution of the individual peaks with the aqueous-organic mixtures was more problematic.