Ionic liquids versus triethylamine as mobile phase additives in the analysis of β-blockers

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Abstract

A comparative study of peak shape, elution behavior, elution strength and resolution of seven β-blockers (acebutolol, alprenolol, labetalol, metoprolol, nadolol, pindolol and propranolol) chromatographed with aqueous-organic mobile phases containing additives such as the ionic liquid (IL) 1-butyl-3-methylimidazolium (BMIM BF4) or triethylamine (TEA) is performed using a conventional reversed-phase Kromasil C18 column and isocratic elution. The efficiencies and asymmetry factors achieved for the group of β-blockers in the Kromasil C18 column improve when the cationic modifiers are added to the aqueous-organic mobile phase as competing additives for the silanol active sites. BMIM BF4 is a significantly better additive compared to TEA. The improvement is more notably for the asymmetry factor, BMIM BF4 allowing to obtain higher plate numbers than TEA at the same concentration. The effects of both modifiers on elution strength and retention factors are, however, different. TEA decreases the solute retention factors when BMIM BF4 does not change them significantly. Using other additives taken in the IL family such as 1-butyl-3-methylimidazolium hexafluorophosphate and 1-octyl-3-methylimidazolium tetrafluoroborate (OMIM BF4), it is shown that the silanol screening effect is always observed, due to the IL cation, when it is possible to increase or to decrease the solute retention factors playing with the hydrophobic nature or chaotropic character of its anion.

Introduction

Octyl (C8) or octadecyl (C18) bonded silica particles are still the most popular chromatographic supports used in columns for reversed phase liquid chromatography (RPLC). The reasons for this success are that these packings are able to separate a wide variety of compounds going from polar, including ionizable, to very apolar compounds. Following the demand, the commercial choice of RPLC C8 or C18 columns is extremely wide [1], [2]. Basic solutes are compounds containing an amino group with pKa higher than 9. In the usual pH range of RPLC mobile phases, 3 < pH < 8, their amine group is positively charged. These amines interact with the residual silanols on the packing material, which leads to some undesirable effects such as asymmetric peaks, low efficiencies and/or irreproducible retention [3], [4]. Many basic compounds have a great interest in the pharmaceutical industry. Various strategies were developed to improve the RPLC analysis of basic compounds reducing or suppressing of the deleterious effects of the amine-silanol strong affinity. The first solution is to suppress the residual silanol groups by changing the bonding chemistry or working with stationary phases non-based on silica [2]. Another solution is to add to the mobile phase silanol masking agents such as amines [5], [6]. The most popular amine is triethylamine (TEA), but cyclohexylamine or dimethyloctylamine were also used successfully [6]. The second solution has the advantage of using the existing equipment explaining that silanol screening agents are actively searched. Recently, room temperature ionic liquids (RTIL) were proposed as possible candidates to reduce or suppress silanol activity [7], [8], [9].

Composed entirely of ions, RTILs are salts with very low melting points, often below room temperature [10]. As salts, they have a very low volatility and flammability [11]. They possess a unique array of physicochemical properties that makes them very attractive solvents in the chemical fields of synthesis, catalysis and electrochemistry [12], [13], [14]. Due to these properties, RTILs start to be employed as promising replacements of organic pollutant solvents [10], [15]. They were consequently considered as benign or green solvents.

The original properties of RTILs were considered for application in the analytical chemistry field. They were used as possible stationary phases in gas chromatography [16], [17]. In their role of silanol blocking agents in RPLC [7], [8], [9], their dual nature should always be kept in mind. Indeed, RTILs are obviously formed by an anion associated to a cation. The chaotropic character of the anion may introduce ion-pairing with cationic solutes and adsorption on the stationary phase. The hydrophobicity of the cation may further induce stationary phase adsorption [18], [19]. It should be noted that when RTILs are used as mobile phase additives at millimolar concentration in the mobile phase, all specific properties of ionic liquids are gone. RTILs become just dissociated salts with hydrophobic cations and chaotropic anions [18].

In a recent work in thin layer chromatography, RTILs of the imidazolium tetrafluoroborate family were demonstrated to be superior to TEA to suppress the deleterious effect of free silanols [20]. The goal of this work is similarly to evaluate the effect of RTILs as silanol screening agents in RPLC and to compare it with TEA. A set of seven basic β-blockers were used as test cationic solutes. β-Blockers are usually administered to treat high blood pressure, cardiovascular, neurological and neuropsychiatric disorders [21]. β-Blockers are also abused in precision sports due to their blood pressure regulation and tremor decreasing effects [21], [22]. The availability of procedures to determine these compounds is mandatory in several fields, including forensic, toxicology and doping control. The seven selected compounds will likely never be administered together to a patient. They were selected as an academic set of cationic solutes with basic properties [27]. TEA was selected as the most popular silanol suppressing agent [6] and 1-butyl-3-methylimidazolium tetrafluoroborate (BMIM BF4) was selected as a typical RTIL easily available. The criteria selected for chromatographic efficiency were of course the peak plate number and the peak asymmetry. The retention factors associated to mobile phase elution strength and the resolution were also measured and compared.

Section snippets

Reagents

Table 1 lists the structures of the seven studied β-blockers along with the literature values of their dissociation constants and molecular and apparent octanol–water partition coefficients at pH 3. All the drugs were purchased from Sigma (L’Isles d’Abeau Chesnes, France). Stock standard solutions (∼100 μg mL−1) of β-blockers were prepared in methanol (SDS Carlo Erba, Peypin, France). Work solutions (∼40 μg mL−1) were conveniently diluted with distilled water. The RTILs 1-butyl-3-methylimidazolium

Peak efficiency and asymmetries

The dissociation constants (pKa) of β-blockers in water are in the 9–10 range (Table 1). It means that they are in their cationic state in the working pH range (3 < pH < 8). The marked hydrophobicity difference between the molecular form of the compound and the cationic form is seen by the 3–4 unit change in respective octanol water partition coefficient listed in Table 1 [23], [24]. For example, the Po/w coefficient of acebutolol in its molecular form is 63 (log Po/w = 1.8); it is reduced to 0.008

Conclusions

Using a conventional C18 column, the efficiencies and asymmetry factors of the peaks corresponding to a group of β-blockers experiment a significant improvement if a cationic competing additive is added to the mobile phase. The ionic liquid BMIM BF4 was found clearly superior to the classical TEA additive for efficiency as well as peak shape enhancement. Furthermore, BMIM BF4 has little influence on the solute retention factors when TEA decreases them. It is shown that the dual nature of the

Acknowledgments

MJRA is grateful to Ministerio de Educación y Ciencia for a Spanish post-doctoral fellowship. SCB thanks the Conselleria d’Empresa, Universitat I Ciencia de la Generalitat Valenciana and AB thanks the Centre National de la Recherche Scientifique (UMR 5180-Lanteri) for French financial support.

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