Electromembrane extraction of peptides--fundamental studies on the supported liquid membrane

A large screening of different components in the supported liquid membrane (SLM) in electromembrane extraction (EME) was performed to test the extraction efficiency on eight model peptides. Electromembrane extraction from a 500 μL acidified aqueous sample containing the model peptides in the concentration 10 μg/mL was used. Extraction time was 5 min with an electric potential of 10 V and 900 rpm agitation of the sample vial. The samples were extracted through a hollow fiber-based SLM with different compositions of organic solvents and carriers. A small volume of acidified acceptor solution (25 μL) was after extraction analyzed directly, or with some dilution, on CE or HPLC. This article has identified mono- or di-substituted phosphate groups as the prominent group of carrier molecules needed to obtain acceptable recoveries. For the organic solvents, primary alcohols and ketones have shown promise regarding recovery and reproducibility, with some differences in selectivity. A new composition of the SLM, namely 2-octanone and tridecyl phosphate (90:10 w/w) has proved to give higher extraction recoveries and lower standard deviation than SLMs previously reported in the literature.     

Impedometric monitoring of the behavior of the supported liquid membrane in electromembrane extraction systems: An insight into the origin of optimized experimental parameters

Electromembrane extraction (EME) was carried out using a novel instrumentation capable of impedometric monitoring of the system during the extraction. This instrumentation involves a classical two-electrode assembly fed by two time-resolved potential functions, the first for the extraction of analyte and the second for obtaining the impedance information. The impedometric analysis of the system was achieved by Laplace transformation of the current recorded during the extraction. It has been shown that the obtained impedance information can be converted to very useful knowledge about time dependence of double layer capacitance, kinetics of analyte depletion, total permeability of the SLM and the effect of experimental parameters on system behavior. It has also been shown that the impedance analysis is a powerful tool for the estimation of optimum experimental parameters without determination of analyte in the acceptor phase.     

Continuous flow liquid membrane extraction: a novel automatic trace-enrichment technique based on continuous flow liquid-liquid extraction combined with supported liquid membrane

Combining the continuous flow liquid-liquid extraction (CFLLE) and supported liquid membrane (SLM) extraction, a novel aqueous-aqueous extraction technique that we termed continuous flow liquid membrane extraction (CFLME) is developed for trace-enrichment. The analyte was firstly extracted into the organic phase in the CFLLE step, then transported onto the organic liquid membrane that formed on the surface of the micro porous membrane of the SLM equipment. Finally, it passed through the liquid membrane and was trapped by the acceptor. Aspects related to CFLME were studied by using dichloromethane as liquid membrane, and sulfonylurea herbicides as model compounds. An enrichment factor of over 1000 was obtained when 10 μg l⁻¹ of MSM was enriched for 120 min by this technique. The drawbacks of only a few organic solvents can be selected as liquid membrane with a limited lifetime in SLM operation was overcome. In this CFLME method, almost all solvents that used in the conventional liquid-liquid extraction (LLE) can be adopted and the lifetime of liquid membrane is no longer a problem.     

Drop-to-drop microextraction across a supported liquid membrane by an electrical field under stagnant conditions

Electromembrane extraction (EME) of basic drugs from 10 μL sample volumes was performed through an organic solvent (2-nitrophenyl octyl ether) immobilized as a supported liquid membrane (SLM) in the pores of a flat polypropylene membrane (25 μm thickness), and into 10 μL 10 mM HCl as the acceptor solution. The driving force for the extractions was 3–20 V d.c. potential sustained over the SLM. The influence of the membrane thickness, extraction time, and voltage was investigated, and a theory for the extraction kinetics is proposed. Pethidine, nortriptyline, methadone, haloperidol, and loperamide were extracted from pure water samples with recoveries ranging between 33% and 47% after only 5 min of operation under totally stagnant conditions. The extraction system was compatible with human urine and plasma samples and provided very efficient sample pretreatment, as acidic, neutral, and polar substances with no distribution into the organic SLM were not extracted across the membrane. Evaluation was performed for human urine, providing linearity in the range 1–20 μg/mL, and repeatability (RSD) in average within 12%.     

Electromembrane extraction of levamisole from human biological fluids

Electromembrane extraction coupled with high-performance liquid chromatography (HPLC) and ultraviolet (UV) detection was developed for the determination of levamisole in some human biological fluids. Levamisole migrated from 4 mL of different acidized biological matrices, through a thin layer of 2-nitrophenyl octyl ether containing 5% tris-(2-ethylhexyl) phosphate immobilized in the pores of a porous hollow fiber, into a 20-μL acidic aqueous acceptor solution present inside the lumen of the fiber. The parameters influencing electromigration were investigated and optimized. Within 15 min of operation at 200 V, levamisole was extracted from different biological fluid samples with recoveries in the range of 59-65%, which corresponded to preconcentration factors in the range of 118-130. The calibration curves showed linearity in the range of 0.5-10, 0.2-10 and 0.1-10 μg/mL for plasma, urine and saliva, respectively. Limits of detection of 0.1, 0.07 and 0.05 μg/mL and limits of quantification of 0.5, 0.2 and 0.1 μg/mL were obtained for plasma, urine and saliva, respectively. The relative standard deviations of the analysis were found to be in the range of 5.6-9.7% (n = 3). Electromembrane extraction was successfully processed for determination of levamisole in plasma, urine and saliva samples.     

Electromembrane extraction of polar basic drugs from plasma with pure bis(2-ethylhexyl) phosphite as supported liquid membrane

Electromembrane extraction (EME) of polar basic drugs from human plasma was investigated for the first time using pure bis(2-ethylhexyl) phosphite (DEHPi) as the supported liquid membrane (SLM). The polar basic drugs metaraminol, benzamidine, sotalol, phenylpropanolamine, ephedrine, and trimethoprim were selected as model analytes, and were extracted from 300 μL of human plasma, through 10 μL of DEHPi as SLM, and into 100 μL of 10 mM formic acid as acceptor solution. The extraction potential across the SLM was 100 V, and extractions were performed for 20 min. After EME, the acceptor solutions were analyzed by high-performance liquid chromatography-ultraviolet detection (HPLC-UV). In contrast to other SLMs reported for polar basic drugs in the literature, the SLM of DEHPi was highly stable in contact with plasma, and the system-current across the SLM was easily kept below 50 μA. Thus, electrolysis in the sample and acceptor solution was kept at an acceptable level with no detrimental consequences. For the polar model analytes, representing a log P range from −0.40 to 1.32, recoveries in the range 25–91% were obtained from human plasma. Strong hydrogen bonding and dipole interactions were probably responsible for efficient transfer of the model analytes into the SLM, and this is the first report on efficient EME of highly polar analytes without using any ionic carrier in the SLM.     

Electromembrane extraction—Three‐phase electrophoresis for future preparative applications

The purpose of this article is to discuss the principle and the future potential for electromembrane extraction (EME). EME was presented in 2006 as a totally new sample preparation technique for ionized target analytes, based on electrokinetic migration across a supported liquid membrane under the influence of an external electrical field. The principle of EME is presented, and typical performance data for EME are discussed. Most work with EME up to date has been performed with low‐molecular weight pharmaceutical substances as model analytes, but the principles of EME should be developed in other directions in the future to fully explore the potential. Recent research in new directions is critically reviewed, with focus on extraction of different types of chemical and biochemical substances, new separation possibilities, new approaches, and challenges related to mass transfer and background current. The intention of this critical review is to give a flavor of EME and to stimulate into more research in the area of EME. Unlike other review articles, the current one is less comprehensive, but put more emphasis on new directions for EME.     

Electromembrane extraction of gonadotropin‐releasing hormone agonists from plasma and wastewater samples

In the present study, for the first time electromembrane extraction followed by high performance liquid chromatography coupled with ultraviolet detection was optimized and validated for quantification of four gonadotropin‐releasing hormone agonist anticancer peptides (alarelin, leuprolide, buserelin and triptorelin) in biological and aqueous samples. The parameters influencing electromigration were investigated and optimized. The membrane consists 95% of 1‐octanol and 5% di‐(2‐ethylhexyl)‐phosphate immobilized in the pores of a hollow fiber. A 20 V electrical field was applied to make the analytes migrate from sample solution with pH 7.0, through the supported liquid membrane into an acidic acceptor solution with pH 1.0 which was located inside the lumen of hollow fiber. Extraction recoveries in the range of 49 and 71% within 15 min extraction time were obtained in different biological matrices which resulted in preconcentration factors in the range of 82–118 and satisfactory repeatability (7.1 < RSD% < 19.8). The method offers good linearity (2.0–1000 ng/mL) with estimation of regression coefficient higher than 0.998. The procedure allows very low detection and quantitation limits of 0.2 and 0.6 ng/mL, respectively. Finally, it was applied to determination and quantification of peptides in human plasma and wastewater samples and satisfactory results were yielded.     

Critical parameters in a supported liquid membrane extraction technique for ionizable organic compounds with a stagnant acceptor phase

The reviews cover important critical parameters that are often optimized in a supported liquid membrane extraction technique in both flat sheet and hollow fibre designs for ionizable organic molecules. Understanding of these parameters can enable one to predict the behavior of the compound before hand and thus reduce the number of optimization experiments. Moreover, less number of experiments can be also generated using statistical techniques which are now becoming more commonly used. Supported liquid membrane extraction optimal parameters such as the conditions of the pH of the acceptor and donor phases should easily be fixed from the pKa values of the compounds. Other parameters, including the polarity of the compound can help to predict the partitioning into the membrane and the behavior of the compound. The influence of parameters such as temperature on the mass transfer in supported liquid membrane depends on the design of the module, experimental design and type of mass transfer controlling the extraction process.     

Electronic simulation of the supported liquid membrane in electromembrane extraction systems: Improvement of the extraction by precise periodical reversing of the field polarity

In order to understand the limitations of electromebrane extraction procedure better, a simple equivalent circuit has been proposed for a supported liquid membrane consisting of a resistor and a low leakage capacitor in series. To verify the equivalent circuit, it was subjected to a simulated periodical polarity changing potential and the resulting time variation of the current was compared with that of a real electromembrane extraction system. The results showed a good agreement between the simulated current patterns and those of the real ones. In order to investigate the impact of various limiting factors, the corresponding values of the equivalent circuit were estimated for a real electromembrane extraction system and were attributed to the physical parameters of the extraction system. A dual charge transfer mechanism was proposed for electromembrane extraction by combining general migration equation and fundamental aspects derived from the simulation. Dual mechanism comprises a current dependent contribution of analyte in total current and could support the possibility of an improvement in performance of an electromembrane extraction by application of an asymmetric polarity changing potential. The optimization of frequency and duty cycle of the asymmetric polarity exchanging potential resulted in a higher recovery (2.17 times greater) in comparison with the conventional electromebrane extraction. The simulation also provided more quantitative approaches toward the investigation of the mechanism of extraction and contribution of different limiting factors in electromembrane extraction. Results showed that the buildup of the double layer is the main limiting factor and the Joule heating has lesser impact on the performance of an electromebrane extraction system.     

Potential‐driven peptide extractions across supported liquid membranes: Investigation of principal operational parameters

Fundamental experiments on electromembrane extraction were performed to increase the basic knowledge about the current and the mass transfer of target peptides and background electrolyte ions. Three peptides (angiotensin 2, bradykinin, and enkephalin) were extracted from 500 μL aqueous donor solution (1 mM HCl, positive electrode), through a 200 μm supported liquid membrane (SLM) of 1‐octanol/di‐isobutylketon/di‐(2‐ethylhexyl) phosphate (55:35:10 w/w/w) sustained in the pores of a porous hollow fiber, and into 25 μL aqueous acceptor solution (50 mM HCl, negative electrode) present inside the lumen of the fiber by the application of an electrical potential (50 V) and agitation (1050 rpm). Recoveries were typically in the range of 55–65% after 5 min of extraction and were principally determined by the chemical composition of the SLM and by the applied voltage. The electrical current in the system was measured during the extraction and was close to 350 μA. The current arose to some extent from mass transfer of the target peptides, but the major contribution was due to a background current from di‐(2‐ethylhexyl) phosphate in the SLM and from mass transfer of background electrolytes. Operation at relatively low background current was important to maintain a stable system.     

In‐line coupling of supported liquid membrane extraction across nanofibrous membrane to capillary electrophoresis for analysis of basic drugs from undiluted body fluids

Planar polyamide 6 nanofibrous membrane was for the first time used in direct coupling of supported liquid membrane (SLM) extraction to CE analysis. Disposable microextraction device with the nanofibrous membrane was preassembled and stored for immediate use. The membrane in the device was impregnated with 1 µL of 1‐ethyl‐2‐nitrobenzene and the device was subsequently filled with 10 µL of acceptor solution (10 mM HCl) and 15 µL of donor solution (sample). The device was in‐line coupled to CE system for selective extraction and direct injection, separation and quantification of model basic drugs (nortriptyline, haloperidol, loperamide and papaverine) from standard saline solutions (150 mM NaCl) and from undiluted human body fluids (urine and blood plasma). Compared to standard polypropylene supporting material, the nanofibrous membrane demonstrated superior characteristics in terms of lower consumption of organic solvents, constant volumes of operational solutions, full transparency and possibility to preassemble the devices. Extraction parameters were better or comparable for the nanofibrous vs. the polypropylene membrane and the hyphenated SLM‐CE method with the nanofibrous membrane was characterized by good repeatability (RSD ≤ 11.3%), linearity (r² ≥ 0.9953; 0.5–20 mg/L), sensitivity (LOD ≤ 0.4 mg/L) and transfer (27–126%) of the basic drugs.     

Fine‐tuning of electromembrane extraction selectivity using 18‐crown‐6 ethers as supported liquid membrane modifiers

Selectivity of electromembrane extractions (EMEs) was fine‐tuned by modifications of supported liquid membrane (SLM) composition using additions of various 18‐crown‐6 ethers into 1‐ethyl‐2‐nitrobenzene. Gradually increased transfer of K⁺, the cation that perfectly fits the cavity of 18‐crown‐6 ethers, was observed for EMEs across SLMs modified with increasing concentrations of 18‐crown‐6 ethers. A SLM containing 1% w/v of dibenzo‐18‐crown‐6 in 1‐ethyl‐2‐nitrobenzene exhibited excellent selectivity for EMEs of K⁺. The established host–guest interactions between crown ether cavities in the SLM and potassium ions in donor solution ensured their almost exhaustive transfer into acceptor solution (extraction recovery ～92%) within 30 min of EME at 50 V. Other inorganic cations were not transferred across the SLM (Ca²⁺ and Mg²⁺) or were transferred negligibly (NH₄⁺, Na⁺; extraction recovery < 2%) and had only subtle effect on EMEs of K⁺. The high selectivity of the tailor‐made SLM holds a great promise for future applications in EMEs since the range of similar selective modifiers is very broad and may be applied in various fields of analytical chemistry.     

Fundamental studies on the electrokinetic transfer of net cationic peptides across supported liquid membranes

By the application of an electrical potential difference (25 V), 37 different peptides were extracted from 500 μL aqueous sample (10 mM formic acid, positive electrode), through a supported liquid membrane (SLM) impregnated in the walls of a porous hollow fiber, and into 25 μL aqueous acceptor solution (100 mM formic acid, negative electrode) present inside the lumen of the fiber. Most of the peptides were obtained by tryptic digestion of cytochrome c and bovine serum albumin, which yielded complex samples for extraction. Three different SLMs were utilized to correlate the peptides extractability with the highly variable physical-chemical properties of the peptides. The first SLM (pure eugenol) provided an electromembrane extraction system for hydrophobic and intermediate peptides (hydrophilicity values below 0.2), where the extraction of peptides into the SLM was mainly based on solvent interactions. The second SLM (1-octanol/di-isobutylketone/di-(2-ethylhexyl) phosphate) extracted both hydrophobic and hydrophilic peptides (hydrophilicity values in the range from -2 to+1) successfully, and the transfer of peptides was principally based on ionic interactions with di-(2-ethylhexyl) phosphate. The third SLM (1-octanol/15-crown-5 ether) was selective for hydrophobic peptides (negative hydrophilicity values), and complexation of the peptides with the crown ether was important for the migration of peptides into the acceptor solution.     

支撑液膜研究及应用进展

支撑液膜技术是高选择性膜分离技术,本文对支撑液膜分离技术的研究进展进行了回顾,详述了用于金属离子分离的支撑液膜所采用的载体、影响支撑液膜稳定性的原因以及改善途径,并对支撑液膜的发展前景进行了展望。     

Simultaneous determination of fluoxetine and its major active metabolite norfluoxetine in human plasma by LC‐MS/MS using supported liquid extraction

A rapid, sensitive and selective bioanalytical method was developed for the simultaneous determination of fluoxetine and its primary metabolite norfluoxetine in human plasma. Sample preparation was based on supported liquid extraction (SLE) using methyl tert‐butyl ether to extract the analytes from human plasma. Chromatography was performed on a Synergi 4 μ polar‐RP column using a fast gradient. The ionization was optimized using ESI (+) and selectivity was achieved by tandem mass spectrometric analysis using MRM functions, m/z 310 → 44 for fluoxetine, m/z 296 → 134 for norfluoxetine and m/z 315 → 44 for fluoxetine‐d₅ (internal standard). The method is linear over the range of 0.05–20 ng/mL (using a human plasma sample volume of 0.1 mL) with a coefficient determination of greater than 0.999. The method is accurate and precise with intra‐batch and inter‐batch accuracy (%bias) of <±15% and precision (%CV) of <15% for both analytes. A run time of 4 min means a high throughput of samples can be achieved. To our knowledge, this method appears to be the most sensitive one reported so far for the quantitation of fluoxetine and norfluoxetine and can be used for routine therapeutic drug monitoring or pharmacokinetic studies. Copyright © 2011 John Wiley & Sons, Ltd.     

Electromembrane extraction of tartrazine from food samples: Effects of nano‐sorbents on membrane performance

In the present study, for the first time electromembrane extraction followed by high‐performance liquid chromatography coupled with ultraviolet detection was developed and validated for the determination of tartrazine in some food samples. The parameters influencing electromembrane extraction were evaluated and optimized. The membrane consists of 1‐octanol immobilized in the pores of a hollow fiber. As a driving force, a 30 V electrical field was applied to make the analyte migrate from sample solution with pH 3, through the supported liquid membrane into an acceptor solution with pH 10. Best preconcentration (enrichment factor >21) was obtained in extraction duration of 15 min. Effects of some solid nano‐sorbents like carbon nanotubes and molecularly imprinted polymers on membrane performance and electromembrane extraction efficiency were evaluated. The method provided the linearity in the range 25–1000 ng/mL for tartrazine (R² > 0.9996) with repeatability range (RSD) between 3.8 and 8.5% (n = 3). The limits of detection and quantitation were 7.5 and 25 ng/mL, respectively. Finally, the method was applied to the determination and quantification of tartrazine from some food samples with relative recoveries in the range between 90 and 98%.     

Electromembrane extraction of basic drugs from untreated human plasma and whole blood under physiological pH conditions

The present work describes the first systematic study of electromembrane extraction (EME) from biological matrices under physiological conditions. Six basic drugs with protein binding in the range of 20–97% were extracted from untreated human plasma and whole blood through a supported liquid membrane (SLM) consisting of 1-ethyl-2-nitrobenzene impregnated in the walls of a hollow fiber, and into an acidified aqueous solution inside the lumen of the fiber. The electrical potential difference over the membrane reduced the protein binding of the drugs and transported the free drug fraction over the membrane. Recoveries in the range 25–65% were obtained with 10-min extraction time and an applied voltage of only 10 V over the SLM. Interday precision better than 20% RSD and linearity in the range 0.5–10 μg/mL were obtained for nortriptyline and methadone. Extraction from untreated whole blood was also demonstrated with recoveries in the range 19–51%.     

Hollow-fibre supported liquid membrane extraction for determination of fluoxetine and norfluoxetine concentration at ultra trace level in sewage samples

In this study, a method was developed for determining the concentration of the pharmaceutical fluoxetine and its metabolite, norfluoxetine, in sewage water samples. Sample preparation was performed by hollow-fibre supported liquid membrane (HF-SLM) extraction with final analysis using liquid chromatography with UV detection. Several parameters were studied including type of organic solvent, sample and acceptor pH, and salt and humic acid content. The optimised method allowed determination of the analyte at the ng/L level in sewage water. A linear plot gave a correlation coefficient better than 0.991 for both analytes and resulted in limits of detection in sewage water of 11 and 12 ng/L, for fluoxetine and norfluoxetine, respectively. The enrichment factor was over 1700 for both analytes in sewage water. The repeatability and reproducibility were better than 8% and 17%, respectively. The developed methodology was used to study daily variations of fluoxetine and norfluoxetine in municipal sewage streams. Norfluoxetine has been detected for the first time in sewage water and a preliminary analysis gave average concentrations of 150 and 225 ng/L for norfluoxetine and fluoxetine, respectively.     

Simultaneous extraction and determination of trace amounts of diclofenac from whole blood using supported liquid membrane microextraction and fast Fourier transform voltammetry

A novel, simple, and inexpensive analytical technique based on flat sheet supported liquid membrane microextraction coupled with fast Fourier transform stripping cyclic voltammetry on a reduced graphene oxide carbon paste electrode was used for the extraction and online determination of diclofenac in whole blood. First, diclofenac was extracted from blood samples using a polytetrafluoroethylene membrane impregnated with 1‐octanol and then into an acceptor solution, subsequently it was oxidized on a carbon paste electrode modified with reduced graphene oxide nanosheets. The optimal values of the key parameters influencing the method were as follows: scan rate, 6 V/s; stripping potential, 200 mV; stripping time, 5 s; pH of the sample solution, 5; pH of the acceptor solution,7; and extraction time, 240 min. The calibration curves were plotted for the whole blood samples and the method was found to have a good linearity within the range of 1–25 μg/mL with a determination coefficient of 0.99. The limits of detection and quantification were 0.1 and 1.0 μg/mL, respectively. Using this coupled method, the extraction and determination were merged into one step. Accordingly, the speed of detection for sensitive determination of diclofenac in complex samples, such as blood, increased considerably.