Microemulsion Liquid Chromatography Research Articles

Methylene selectivity in microemulsion liquid chromatography

An example of the chromatographic analysis of a mixture of benzene homologues in a microemulsion liquid chromatography (MELC) mode with different surfactants shows that the dependence of the retention of the number of methylene groups has a convex character that is fundamentally different from the normal mode of reversed phase (RP) high performance liquid chromatography (HPLC), where the relationship is exponential. The mechanism of sorption of analytes in MELC is proposed. The analysis of a test mixture of alkylbenzenes shows a difference in the behavior of the enthalpy of sorption in MELC and HPLC modes.

Microemulsion liquid chromatographic method for simultaneous separation and determination of six flavonoids of Apocynum venetum leaf extract.

A simple, cost-effective, and efficient method was developed for the rapid simultaneous separation and determination of six flavonoids (rutin, hyperoside, quercetin-3-O-sophoroside, isoquercitrin, astragalin and quercetin) of Apocynum venetum leaf extract by reversed phase high performance liquid chromatography using a microemulsion system mixture as the mobile phase. Separations were performed on the Zorbax Extend-C18 column with UV detection at 360nm. The flow rate was 0.8mLmin(-1). The optimized microemulsion mobile phase consisted of 2.5% (v/v) n-butanol, 1.2% (v/v) of Genapol X-080, 0.5% (v/v) ethyl acetate and 95.8% (w/v) of aqueous 20mM phosphoric acid, pH adjusted to 6.0 with 0.3% triethylamine. Under the optimized conditions, the calibration curve for six flavonoids was linear in the range of 5-1000μgmL(-1) with the correlation coefficients greater than 0.9994. The intra-day and inter-day precision (RSD) were below 8.11% and the limits of detection (LOD) for the six flavonoids were 1.7-6.0μgmL(-1) (S/N=3). The microemulsion liquid chromatography (MELC) method was successfully applied to separate and determine the six flavonoids of A. venetum leaf extract.

Determination of drug lipophilicity by phosphatidylcholine-modified microemulsion high-performance liquid chromatography

A new biomembrane-mimetic liquid chromatographic method using a C8 stationary phase and phosphatidylcholine-modified (PC-modified) microemulsion mobile phase was used to estimate unionized and ionized drugs lipophilicity expressed as an n-octanol/water partition coefficient (logP and logD). The introduction of PC into sodium dodecyl sulfate (SDS) microemulsion yielded a good correlation between logk and logD (R2=0.8). The optimal composition of the PC-modified microemulsion liquid chromatography (PC-modified MELC) mobile phase was 0.2% PC—3.0% SDS—6.0% n-butanol—0.8% ethyl acetate—90.0% water (pH 7.0) for neutral and ionized molecules. The interactions between the analytes and system described by this chromatographic method is more similar to biological membrane than the n-octanol/water partition system. The result in this paper suggests that PC-modified MELC can serve as a possible alternative to the shake-flask method for high-throughput unionized and ionized drugs lipophilicity determination and simulation of biological processes.

Determination of the lipophilicity (log Po/w) of organic compounds by microemulsion liquid chromatography

Four microemulsion liquid chromatography (MELC) systems and one micellar liquid chromatography (MLC) system have been evaluated as potential high-throughput screening platforms capable of modeling the partitioning behaviors of drug compounds in an n-octanol/water system and predicting their lipophilicity (i.e., logP values). The microemulsion mobile phases is consisted of sodium dodecyl sulfate (SDS), butanol, octane, heptanes, octanol and water. A linear solvation energy relationship (LSER)-based method was used to compare the MELC and MLC systems, as well as several other biochemical systems, and to identify the optimal system by comparing their Euclidean distances with the LSER coefficients. The most effective MELC system had a mobile phase consisting of 3.0% (w/w) SDS, 6.0% (w/w) butanol, 0.8% (w/w) octanol, and 90.2% (w/w) water (pH 6.4). The results showed that it gave superior results to the other chromatographic systems in terms of its ability to predict the logP values of drug compounds.

Investigation of influence of nonionic additives on structural changes of water droplets encapsulated in AOT reverse micelles by instrumental methods

The microstructure of sodium bis(2-ethylhexyl) sulfosuccinate reverse micelles was investigated using infrared, nuclear magnetic resonance and ultraviolet–visible spectroscopic methods, also by water-in-oil microemulsion chromatography. The influence of structure-making and structure-breaking nonionic additives on the microenvironment in reverse micelles was studied. The values of chemical shift in the presence of chaotropic urea are different than their values in case of kosmotropic glucose additives according to proton magnetic resonance spectroscopic data. Urea promotes an increasing of free water fraction in the water pockets of the reverse microemulsions, but glucose supports formation of the trapped water on the basis of deconvolution of the OH stretching vibrational absorption spectra in the region of 3000–3800cm−1 into three subpeaks with a Monte Carlo method. The different influence of kosmotropic and chaotropic nonionic additives on both binding constant Kb of o-nitroaniline to the micelles of AOT and retention of the model compounds were also revealed by UV–visible spectroscopic and water-in-oil microemulsion liquid chromatography methods respectively.

Optimization of liquid chromatographic method for the separation of nine hydrophilic and hydrophobic components in Salviae miltiorrhizae Radix et Rhizoma (Danshen) using microemulsion as eluent

In this study, we have proposed and developed a novel, environmental-friendly and simple method for separation of nine hydrophilic and hydrophobic components in Danshen using microemulsion liquid chromatography. The proposed method was optimized via the preliminary screening experiment and the experimental design. The following factors were investigated in preliminary screening experiment: pH of mobile phase, column type, the nature of surfactant, the nature of oil phase and additives. In order to simultaneously optimize resolution and analysis time, the chromatographic optimization function (COF) was adopted to evaluate chromatograms. The central composite design (CCD) was used to create the matrix of experiments for mapping the chromatographic response surface. Finally, the COF values were fitted into a second order polynomial model and the response surface methodology (RSM) was employed to find the optimal eluent constituents. The reliability of the established model was confirmed by the good agreement obtained between experimental data and predictive values. Based on the results from the preliminary screening experiment and the CCD optimization, the optimal mobile phase was identified as a solution consisting of 6.68% (w/w) polyoxyethylene lauryl ether (Brij35), 0.84% (w/w) cyclohexane, 6.92% (w/w) n-butanol, 85.56% (w/w) phosphate buffer (pH 6.60) and 8mM cetyltrimethyl ammonium bromide (CTAB).

Simultaneous separation and determination of four phenylethanoid glycosides in rat plasma sample after oral administration of Cistanche salsa extract by microemulsion liquid chromatography

A simple, rapid and specific method was developed to separate as well as to determine the four phenylethanoid glycosides (PhGs) (echinacoside, tubuloside B, acteoside and isoacteoside) in rat plasma after oral administration of Cistanche salsa extract by reversed phase high performance liquid chromatography using a microemulsion as the mobile phase. The separations were performed on a Zorbax Extend-C18 column at 25°C. Photodiode-array detector was conducted at 322nm and with a flow rate of 0.8mLmin(-1). The optimized microemulsion mobile phase consisted of 0.3% triethylamine in 20mM phosphoric acid at pH 6.0, 0.8% (v/v) ethyl acetate as oil phase, 1.5% (v/v) Genapol X-080 as surfactant, 2.5% (v/v) n-propanol as co-surfactant. Under the optimal conditions, the calibration curve for four PhGs was linear in the range of 10-1000ngmL(-1) with the correlation coefficients greater than 0.9994. The intra-day and inter-day precision (RSD) were below 8.64% and the limits of detection (LOD) for the four PhGs were 0.4-1.3ngmL(-1) (S/N=3). The microemulsion liquid chromatography (MELC) method was successfully applied to separate and determine the four PhGs in rat plasma after oral administration of C. salsa extract.

Determination of three phenolic compounds in water samples by solid phase extraction with microemulsion liquid chromatography

A method for the determination of phenol, bisphenol A (BPA), 2,4-dichlorophenol in water samples based on solid-phase extraction with microemulsion liquid chromatography (MELC) was established. The water sample was acidified, and then cleaned-up by a C18 solid phase extraction cartridge. The separation was carried out on an Inertsil C18 (150 mm x 4.6 mm, 5 microm) with a gradient elution using a microemulsion (consisting of 3.0% sodium dodecyl sulfate (SDS), 6.0% butyl alcohol, 0.8% n-heptane, 90.2% (water + 0.5% acetic acid)) and acetonitrile as mobile phases at a flow rate of 1.0 mL/min and a detection wavelength of 280 nm. The detection limits (S/N = 3) of phenol, BPA, 2,4-dichlorophenol were 0.74, 8.0, 8.0 microg/L, respectively. Good linearities were achieved for the target compounds over the range of 0.1 - 10 mg/L. The spiked recoveries of three phenolic compounds in blank water samples) were 82.7%, 87.8%, 82.6% for phenol, BPA, 2,4-dichlorophenol respectively. The recoveries of target compounds in real water samples ranged from 85.7% to 113.2%. The developed method is simple, selective, sensitive and applicable for the analysis of environmental water samples.

Desirability-based optimization and its sensitivity analysis for the perindopril and its impurities analysis in a microemulsion LC system

In the current paper the application of multiobjective optimization (MOOP) technique, via Derringer's desirability function, to a microemulsion liquid chromatographic (MELC) method is described. Chromatographic separation of perindopril tert-butylamine and its four impurities was selected as the case study. Central composite design (CCD) with fractional factorial design, ±0.5 α star design and four replications in central point was applied for a response surface study, in order to examine in depth the effects of the most important factors. As factors that influence the system mostly (i) content of ethyl acetate and (ii) butyl acetate in composite internal phase, (iii) content of sodium dodecyl sulfate (surfactant) and (iv) n-butanol (co-surfactant), as well as (v) pH of the mobile phase were selected. Retention factor of (a) perindoprilat and (b) impurity Y 31 and (c) resolution factor for impurities Y 32 and 33 were chosen for simultaneous optimization. By adjustment of the importance coefficients and weights, according to defined objectives, the optimal mobile phase composition was predicted to be: 0.24% w/v butyl acetate, 0.3% w/v ethyl acetate, 2% w/v SDS, 7.75% w/v n-butanol and pH of the mobile phase 3.7. The sensitivity analysis of desirability function for these optimal conditions was conducted for the first time in LC separations, by applying a sensitivity procedure. The performed sensitivity analysis confirmed that the higher overall desirability does not necessarily mean a better solution. The accuracy of prediction might be affected if the optimal levels of input variables, achieved from several design points, end up with equal settings and different corresponding overall desirability. In our study this was not the issue, which confirmed the adequacy of predicted optimum.

Prediction of polar surface area of drug molecules: A QSPR approach

A quantitative structure-property relationship (QSPR) study based on an artificial neural network (ANN) was carried out for the prediction of the microemulsion liquid chromatography polar surface area (PSA) of a set of 32 drug compounds. The genetic algorithm-kernel partial least squares (GA-KPLS) method was used as a variable selection tool. A KPLS method was used to select the best descriptors and the selected descriptors were used as input neurons in neural network model. For choosing the best predictive model from among comparable models, square correlation coefficient Q(2) for the whole set calculated based on leave-group-out predicted values of the training set and model-derived predicted values for the test set compounds is suggested to be a good criterion. Finally, to improve the results, structure-property relationships were followed by nonlinear approach using artificial neural networks and consequently better results were obtained. Also this demonstrates the advantages of ANN.

Validation of an Oil-in-Water Microemulsion Liquid Chromatography Method for Analysis of Perindopril tert-Butylamine and Its Impurities

This paper describes the development and validation of a microemulsion liquid chromatography (MELC) method for simultaneous determination of perindopril tert-butylamine and its impurities in bulk active substances and the pharmaceutical dosage form of tablets. An appropriate resolution with reasonable retention times was obtained for a microemulsion containing 0.24% (w/v) butyl acetate, 0.30% (w/v) ethyl acetate, 2% (w/v) sodium dodecyl sulfate, 7.75% (w/v) n-butanol, and 20.0 mM potassium dihydrogen phosphate, the pH of which was adjusted to 3.70 with 85% orthophosphoric acid. Separations were performed on a Nucleosil 120-5 butyl modified (C4), 250 x 4 mm, 5 microm particle size silica column at 40 degrees C, with a mobile phase flow rate of 1.25 mL/min. UV detection was performed at 254 nm. The established method was subjected to method validation, and required validation parameters were defined. Robustness testing, an important part of method validation, was performed as well. Since robustness validation can be conducted using different experimental designs, the Plackett-Burman design was applied due to its possibility of testing many factors at the same time. The validated MELC method was found to be suitable for the simultaneous determination of perindopril tert-butylamine and its impurities in pharmaceuticals.

Retention behavior of statins in microemulsion liquid chromatography

The effects of the concentration of surfactant, co-surfactant and lipophilic organic solvent and the pH value of mobile phase on the retention behavior of statins in microemulsion liquid chromatography have been investigated using pravastatin sodium, atorvastatin calcium, lovastatin and simvastatin as model compounds. The experimental results showed that the effects of the concentration of surfactant, co-surfactant and lipophilic organic solvent on the retention behavior of statins in microemulsion liquid chromatography were fully consistent with the theoretical modeling. The effect of the pH value of the mobile phase on the retention behavior of acid statins was fully consistent with the theoretical modeling. The relationship between the retention factor of neutral statins and the concentration of hydrogen ion was an implicit function. The retention modeling can reflect the effect of composition of microemulsion solution on the retention behavior of statins drugs.

Quantitative determination of capsaicin in liniments by microemulsion liquid chromatography

A technique for the quantitative determination of capsaicin in liniments by the microemulsion liquid chromatography method is proposed. Due to the absence of the homogenization, extraction and filtration stages, the procedure of sample pretreatment takes 5 minutes, which is much more rapid in comparison with the variants described in the literature (from 30 minutes to 12 hours). The full time of the analysis (including the procedure for sample pretreatment) is 20 minutes. The graduation curve is linear in the range from 0.02 to 240 mg/l. The limit of determination of capsaicin was 8 μg/I.

Retention Behavior of Acid Drugs in Microemulsion Liquid Chromatography

Retention Behavior of Acid Drugs in Microemulsion Liquid Chromatography

Simultaneous microemulsion liquid chromatographic analysis of fat-soluble vitamins in pharmaceutical formulations: Optimization using genetic algorithm

An environmentally benign and simple method has been proposed for separation and determination of fat-soluble vitamins using isocratic microemulsion liquid chromatography. Optimization of parameters affecting the separation selectivity and efficiency including surfactant concentration, percent of cosurfactant (1-butanol), and percent of organic oily solvent (diethyl ether), temperature and pH were performed simultaneously using genetic algorithm method. A new software package, MLR-GA, was developed for this purpose. The results indicated that 73.6 mM sodium dodecyl sulfate, 13.64% (v/v) 1-butanol, 0.48% (v/v) diethyl ether, column temperature of 32.5 °C and 0.02 M phosphate buffer of pH 6.99 are the best conditions for separation of fat-soluble vitamins. At the optimized conditions, the calibration plots for the vitamins were obtained and detection limits (1.06–3.69 μg mL −1), accuracy (recoveries > 94.3), precision (RSD < 3.96) and linearity (0.01–10 mg mL −1) were estimated. Finally, the amount of vitamins in multivitamin syrup and a sample of fish oil capsule were determined. The results showed a good agreement with those reported by manufactures.

FORCED DEGRADATION STUDIES OF SIMVASTATIN USING MICROEMULSION LIQUID CHROMATOGRAPHY

In this paper, the definition of simvastatin degradation profile by microemulsion liquid chromatography (MELC) is presented. The aim of the study was to investigate simvastatin stability after various stress tests, such as: acid and base hydrolysis, oxidation, and heat. The kinetics of acid and oxidative degradation was also studied. The complex retention mechanism occurring when microemulsion is used as eluent enabled the successful separation of a large number of degradants in prepared stress samples. The existence of a second partitioning site due to microstructural and interfacial characteristics of o/w microemulsion eluent facilitated HPLC separation. All the expected capabilities of MELC method were realized, demonstrating the advantage and value of the MELC method for a wide area of pharmaceutical analysis. For acid degradation the second order rate constant and half-life were calculated. Oxidative decomposition proved to be the first order reaction for which the rate constant and half-life were also determined.

Robustness Testing of Microemulsion Liquid Chromatographic Separation of Simvastatin and its Impurities

Liquid chromatography employing the microemulsion as eluent was applied for the analysis of simvastatin and its six impurities. Previously, the optimized and validated method was tested to prove the method's capability to perform during the small changes of important parameters. In this paper, influence of parameters tipical for microemulsion liquid chromatography (MELC) on the method's robustness was presented. For the results evaluation, and presentation, two statistical methods were applied. In order to designate the most convinient one, multiple regression (MR) and artificial neural networks (ANN) were applied and compared. A set of experiments was defined by the central composite design (CCD). Indepedent variables (SDS, n-buthanol and diisopropyl ether content) and dependent variables (retention factors of investigated substances) were the framework for both, MR and ANN. This study demonstrated that MR and radial basis function ANN are useful tools in understanding the effects of the investigated factors on the chromatographic system and definition of the robustness limits.

Microemulsion liquid chromatographic screening of Simvastatin and its active metabolite in human plasma

Summary This paper reports development and validation of a new microemulsion liquid chromatographic (MELC) method for rapid screening of simvastatin and simvastatin acid in human plasma. Plasma samples were injected directly into the HPLC system after appropriate sample dilution with mobile phase. Separations were performed on a 4.6 mm × 150 mm, 5-μm particle, C18 column, with UV detection at 238 nm. The mobile phase was 0.5% (w/υ) diisopropyl ether, 1.0% (w/υ) sodium dodecylsulphate (SDS), 4.0% (w/v) n-butanol, and 94.5% (w/w) aqueous 25 mM disodium hydrogen phosphate, pH 7.0, at a flow rate of 1 mL min−1. The method was evaluated according to criteria stated in FDA bioanalytical method validation guidance. The unique approach applied in this paper enables direct analysis of simvastatin and simvastatin acid, so the method can be used to obtain reliable results in a rapid and simple way.

Predicting blood–brain barrier penetration of drugs by microemulsion liquid chromatography with corrected retention factor

This study attempted to optimize the compositions of microemulsion mobile phase to model the drug penetration across blood–brain barrier (BBB) by microemulsion liquid chromatography (MELC). During the study, a continuous increase of the retentions along the operation time was found for all the test drugs. A correction method using methyl paraben and propranolol as the internal standards was proposed, which improved the reliability of the retention time significantly. The corrected retention factors were then used to develop the predictive model. The optimized MELC system with microemulsion consisting of 3.3% sodium dodecyl sulfate (SDS)–6.6% butanol–1.6% heptane–88.5% phosphate buffer (pH 7.0) showed superiorities to other lipophilicity measuring systems for modeling BBB penetration.

Characterization of microemulsion liquid chromatography systems by solvation parameter model and comparison with other physicochemical and biological processes

The solvation parameter model has been applied to characterize four microemulsion liquid chromatography (MELC) systems and two micellar liquid chromatography (MLC) systems, and utilized to compare the above systems with other physicochemical and biological processes in this study. The microemulsion mobile phases were composed of sodium dodecyl sulfate (SDS), polyoxyethylene (23) lauryl ether (Brij 35), butanol, heptane and phosphate buffer (pH 7.0) at the designated ratios. The results showed the main difference between the concerned MELC and MLC systems was the decrease of hydrogen-bond basicity of stationary phase with the addition of heptane in microemulsion. Principal component analysis with normalized coefficients can provide consistent results involving the similarities among various systems with that obtained by distance parameter d. Except for some proven similarities of chromatographic systems to octanol–water partition coefficients (log P) and human skin permeation (log K p), a microemulsion HPLC system, the mobile phase being 3.3% SDS–6.6% butanol–1.6% heptane–88.5% buffer, was found very similar to drug penetration across blood–brain barrier and its predictive capability for this biological process was originally evaluated in this study.