Use Of Micellar Mobile Phases Research Articles

Use of micellar mobile phases for the chromatographic determination of melamine in dietetic supplements

Melamine is a nitrogen-rich industrial chemical which is occasionally used to increase the apparent protein content of different products destined for human and animal consumption. In this work, a liquid chromatographic procedure that uses micellar mobile phases of sodium dodecyl sulfate (SDS) buffered at pH 3, a C18 column and UV detection is reported for the determination of melamine in dietetic supplements. Samples were reconstituted with a SDS solution and were directly injected, thus avoiding long extraction and experimental procedures. Melamine was eluted in less than 10 min with no interference by other compounds of the matrices. The optimum mobile phase composition was taken by a chemometrical approach that considers the retention factor, efficiency and peak shape. Validation was performed following the indications of the European Commission (Decision 2002/657/EC). The following parameters were considered: linearity (0.02-100 μg mL(-1); R(2) = 0.9996), intra- and inter-day precisions (<12.4%), accuracy (90.0-101.3%), and robustness (less than 9.8% and 5.1%, for retention time and peak area, respectively). The limits of detection and quantification were 9 and 20 ng mL(-1), respectively. Recoveries for several spiked samples were in the 85.8-114.3% range. These results indicate that the proposed methodology is useful for routine analysis of control quality of infant formula and adult dietetic supplements.

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.

Use of micellar mobile phases for the chromatographic determination of clorazepate, diazepam, and diltiazem in pharmaceuticals.

An ODS-2 column, a micellar mobile phase of high elution strength containing 0.1M sodium dodecyl sulfate and 3% (v/v) butanol, and ultraviolet detection at 230 nm are used for the determination of either of two benzodiazepines (clorazepate and diazepam) and a benzothiazepine (diltiazem) in pharmaceuticals. The procedure is shown to be competitive against conventional chromatography with methanol-water mobile phases, especially for diltiazem. The composition of the micellar mobile phase is selected using a predictive strategy based on an accurate retention model and assisted by computer simulation. Calibration graphs are linear at least in the 2.5 to 20 microg/mL, 4 to 20 microg/mL, and 5 to 40 microg/mL ranges for clorazepate, diazepam, and diltiazem, respectively. The intra- and interday repeatabilities (%) are clorazepate (1.7, 5.2), diazepam (0.43, 3.7), and diltiazem (0.36, 3.1). Limits of detection are well below the concentrations of the drugs found in the commercial pharmaceutical preparations analyzed. The drug contents evaluated with the proposed procedure are compared with the declared contents given by the manufacturers. The achieved percentages of label claim are usually between 95 and 104%.

Development and Validation of a Procedure for Estimating the Hydrophobicity of Structurally Unrelated Compounds by Micellar Liquid Chromatography

Reversed-phase liquid chromatography has been used most often to estimate values of log P, but despite years of study, there is no universally accepted method of performing these estimations. The main problem has to do with the fact that the hydrophobic parameter, log k w , depends on the hydrogen bond acceptor-donor character of the compounds. The use of micellar mobile phases to perform these estimations is evaluated here, and the influence of the nature of the surfactant (anionic, cationic, and nonionic) on the log k-log P relationships is studied. The use of a nonionic surfactant, such as Brij35, to prepare the mobile phases provided adequate results regardeless of the hydrogen bond acceptor-donor character of the compounds, whereas noncongener behaviors are found with anionic and cationic surfactants. This enabled the establishment of a calibration set consisting of 7 compounds with variable hydrophobicity in order to calibrate new columns and predict log Pvalues. In these conditions, the hydrophobicity of structurally unrelated compounds in the -0.1 < log P < 4.5 range can be measured. To evaluate the accuracy of predictions, the hydrophobicity of 27 compounds is determined and compared with the log P values calculated using the ACD-log P program. Nonsignificant differences between the predicted and the theoretical log P values were achieved at 95% probability level

Chromatographic quantitation of the hydrophobicity of ionic compounds by the use of micellar mobile phases

Many biologically active compounds of interest in structure–activity relationships are ionic at physiological pH. However, ionic organic compounds are only weakly or not retained in conventional RPLC which impedes the chromatographic estimation of their hydrophobicity and the development of quantitative retention–activity relationship studies. The use of micellar mobile phases allows the retention of ionic compounds. Hydrophobic and electrostatic forces govern the retention of ionic compounds in micellar liquid chromatography. In this paper three different retention models log k–log P for ionic compounds are tested ( P=partition coefficient). The retention model (log k= a log P+ b α+ c) which includes the hydrophobicity and the molar total charge of compound at a given pH value has proven to be valid for all types of compounds tested, catecholamines, local anesthetics, diuretics and o-phthalaldehyde–N-acetyl- l-cysteine amino acid derivatives.

Quantitative retention–structure and retention–activity relationships of barbiturates by micellar liquid chromatography

Studies on the structural requirements of chromatographic surfaces to emulate in vitro the partitioning process in biomembranes are of great interest. The use of micellar mobile phases in RPLC modifies the hydrophobicity of the stationary phase and provides hydrophobic and electrostatic sites of interaction as a consequence of the adsorption of surfactant monomers to the chromatographic surface. Modified stationary phases in MLC could be structurally similar to biomembranes, but thorough studies are necessary to confirm this. In this paper we focus our attention on barbiturates. The influence of the nature and concentration of the surfactant (Brij 35, SDS and CTAB) and the mobile phase pH on the retention of 13 barbiturates in modified C 18 stationary phases is studied. Quantitative structure–retention and structure–activity relationships for the barbiturates with different surfactants are proposed and compared with those obtained using hydro–organic mobile phases.

Use of Micellar Mobile Phases and an HPLC Column Switching System for Direct Injection Determination of Urinary Free Cortisol

Abstract A direct injection method for the determination of urinary free Cortisol by an HPLC column switching system is presented. The first chromatographic column, which provides for sample preconcentration and clean-up, employs a micellar mobile phase. The second column, coupled on -line to the first, utilizes reversed-phase condition for analysis. UV detection is employed at 254 nm. Detection limit for urinary free Cortisol is 2.4 ng when 2 ml urine sample is injected. Nine samples were analyzed. The results were in a reasonable range. This method can be used successfully in the diagnosis of patients with hypercortisolism and Cushing's syndrome.

Modelling of the retention behaviour of solutes in micellar liquid chromatography with organic modifiers

Most of the reported procedures for the determination of compounds by micellar liquid chromatography make use of micellar mobile phases containing an alcohol. The retention of a solute in a purely micellar eluent has been adequately described by the linear equation 1/ k′ vs. micelle concentration. This equation seems also to be valid for mobile phases with the same alcohol concentration and varying micelle concentrations. A model to describe the retention behaviour of solutes in any mobile phase of surfactant and alcohol is proposed, which makes use of the elution data in five mobile phases of surfactant with different amounts of alcohol. A function of the type 1/ k′ = Aμ + Bϕ + Cμϕ + D, where μ and ϕ are surfactant and alcohol concentration, respectively, proved to be satisfactory for different solutes (catecholamines, amino acids, phenols and other aromatic compounds).

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.

Use of micellar mobile phases and microbore column switching for the assay of drugs in physiological fluids

The feasibility of directly assaying drugs in physiological fluids using on-line preconcentration and microbore high-performance liquid chromatography has been demonstrated. The untreated sample is injected onto a hydrophobic pre-column, using micellar sodium dodecyl sulfate (SDS) in the case of serum or phosphate buffer in the case of urine, as the load mobile phase. This traps the components of interest which are then backflushed onto a microbore analytical column using a stronger mobile phase. This procedure was then applied to diazepam in serum and phenobarbital in urine. Recovery was linear and quantitative over the range 30–3000 ng/ml for diazepam in serum and 2–200 μg/ml for phenobarbital in urine. The diazepam method was specific against caffeine and the three major metabolites of diazepam: oxazepam, temazepam, and nordiazepam. The effects of varying pre-column dimensions, pre-column loading time, and SDS concentration volume were evaluated.

Direct injection of plasma into column liquid chromatographic systems

Since the late 1970’s several techniques involving the direct injection of biological fluids, like plasma and serum have been published. Most of the systems presented utilize a precolumn for the reception of the biological fluids, trace enrichment and preliminary clean-up before the analyte fraction is transferred to the separation column. The most common solid phase for the precolumn is silanized silica, but many other types have been used. One was especially designed for the purpose of direct injections of biological fluids: the Internal Surface Reversed Phase support. The main problem with direct injections is the development of column instabilities due to the denaturation and precipitation of plasma proteins on the support. It has, however, been possible to design systems which can handle the injection of 25ml total plasma volume without the need to exchange precolumn. The most promising techniques in this respect are the so called “pre-column” venting plug technique and the use of micellar mobile phases. Losses of analytes may be a problem when protein-binding extends 95%. However, remedies are available, such as the use of more hydrophobic solid phases, the addition of competitive binders and pH adjustments of injected solutions.

Retention behavior of triglycerides on reverse phase columns using pseudophase liquid chromatography

The use of micellar mobile phases in decreasing the elution time of peanut triglycerides on 10µ reverse phase columns is demonstrated. Both cationic and anionic surfactants were used with “end-capped” and “non-end-capped” reverse phase columns. Surfactants added to both nonaqueous and aqueous organic mobile phases reduced the elution time of all triglyceride peaks when compared to a mobile phase without a surfactant modifier. Sodium dodecyl sulfate (SDS) as the modifier in the mobile phase was the most effective in reducing elution time for triglycerides when an “end-capped” column was used as the stationary medium. Increasing concentrations of surfactant in the mobile phase resulted in corresponding decreases in elution times. A unique feature of micellar phases is that any increase in concentration only alters the concentration of the micelles in the mobile phase. The objective of this study was to investigate the retention behavior of large, non-ionic solute molecules and to elucidate the adsorption mechanism on reverse phase columns.