Optimization of generic conditions for electromembrane extraction of basic substances of moderate or low polarity.

Generic electromembrane extraction (EME) methods were developed and optimized for basic analytes of moderate or low polarity, employing prototype conductive vial EME equipment. Two generic methods, B1 and B2, were devised for mono- and dibasic compounds with distinct polarity windows: 2.0 < log P < 6.0 for B1 and 1.0 < log P < 4.5 for B2. In B1, 10 µL of 2-nitrophenyl octyl ether served as the liquid membrane, while B2 utilized 10 µL of 2-undecanone. Both methods involved the acidification of 125 µL of human plasma samples with 125 µL of sample diluent (0.5 M HCOOH for B1 and 1.0 M HCOOH for B2). The acceptor phase consisted of 250 µL of 100 mM HCOOH. Extraction was conducted for 30 min with agitation at 800 rpm, employing an extraction potential of 100 V for B1 and 50 V for B2. A set of 90 pharmaceutical compounds was employed as model analytes. Both B1 and B2 demonstrated high recoveries (40%-100%) for the majority of model analytes within their respective polarity windows. Intra-day precision was within 2.2% and 9.7% relative standard deviation. Both extraction systems exhibited stability in terms of current, matrix effect values were between 90% and 109%.

Generic Liquid Membranes for Electromembrane Extraction of Bases with Low or Moderate Hydrophilicity.

For the first time, this paper introduces the idea of generic extraction conditions in electromembrane extraction (EME), where the selection of the liquid membrane is based on the charge (z) and hydrophobicity (log P) of the analyte. A broad range of organic solvents were tested as liquid membranes, and 90 basic pharmaceuticals were used as model analytes (-4.2 < log P < 8.1). 2-Nitrophenyl octyl ether (NPOE) was confirmed as a highly efficient liquid membrane for mono- and dibases (+1.0 ≤ z ≤ +2.0) of low polarity in the log P range of 2.2-6.4. This log P range was set as the extraction window (operational range) of NPOE. NPOE provided very high operational stability. At 50 V, the current was at a 1 µA level, and gas formation and drifting pH due to electrolysis were insignificant. 2-Undecanone was discovered as a new and robust alternative. This solvent extracted monobasic analytes (z = +1) in the log P range of 1.0-5.8 and was efficient even for bases of moderate polarity. The current was at the 1-3 µA level when 2-undecanone was operated at 50 V. Tri(pentyl) phosphate emerged as another new alternative for bases in the log P range of 0.5 to 5.5, providing greater selectivity differences. This solvent provided a higher current (30-50 µA), but the EME system stability was not compromised. 2-Undecanone and tri(pentyl) phosphate extracted protonated bases mainly by hydrogen bond interactions. NPOE, on the other hand, extracted based on a combination of hydrogen bond and π-type interactions and was consequently less selective.

Conductive vial electromembrane extraction of opioids from oral fluid.

The use of oral fluid as sample matrix has gained significance in the analysis of drugs of abuse due to its non-invasive nature. In this study, the 13 opioids morphine, oxycodone, codeine, O-desmethyl tramadol, ethylmorphine, tramadol, pethidine, ketobemidone, buprenorphine, fentanyl, cyclopropylfentanyl, etonitazepyne, and methadone were extracted from oral fluid using electromembrane extraction based on conductive vials prior to analysis with ultra-high performance liquid chromatography-tandem mass spectrometry. Oral fluid was collected using Quantisal collection kits. By applying voltage, target analytes were extracted from oral fluid samples diluted with 0.1% formic acid, across a liquid membrane and into a 300 µL 0.1% (v/v) formic acid solution. The liquid membrane comprised 8 µL membrane solvent immobilized in the pores of a flat porous polypropylene membrane. The membrane solvent was a mixture of 6-methylcoumarin, thymol, and 2-nitrophenyloctyl ether. The composition of the membrane solvent was found to be the most important parameter to achieve simultaneous extraction of all target opioids, which had predicted log P values in the range from 0.7 to 5.0. The method was validated in accordance to the guidelines by the European Medical Agency with satisfactory results. Intra- and inter-day precision and bias were within guideline limits of ± 15% for 12 of 13 compounds. Extraction recoveries ranged from 39 to 104% (CV ≤ 23%). Internal standard normalized matrix effects were in the range from 88 to 103% (CV ≤ 5%). Quantitative results of authentic oral fluid samples were in accordance with a routine screening method, and external quality control samples for both hydrophilic and lipophilic compounds were within acceptable limits.

Electromembrane extraction in microfluidic formats.

Electromembrane extraction is a microextraction technique where charged analytes are extracted across a supported liquid membrane and selectively isolated from the sample based on an electrical field. Since the introduction in 2006, there has been continuously increasing interest in electromembrane extraction, and currently close to 50 new articles are published per year. Electromembrane extraction can be performed in different technical configurations, based on standard laboratory glass vials or 96-well plate systems, and applications are typically related to pharmaceutical, environmental, and food and beverages analysis. In addition to this, conceptual research has developed electromembrane extraction into different milli- and microfluidic formats. These are much more early-stage activities, but applications among others related to organ-on-chip systems and smartphone detection indicate unique perspectives. To stimulate more research in this direction, the current article reviews the scientific literature on electromembrane extraction in milli- and microfluidic formats. About 20 original research articles have been published on this subject so far, and these are discussed critically in the following. Based on this and the authors own experiences with the topic, we discuss perspectives, challenges, and future research.

Versatile Integration of Liquid-Phase Microextraction and Fluorescent Aptamer Beacons: A Synergistic Effect for Bioanalysis.

Fluorescent aptamer beacons (FABs) are a major category of biosensors widely used in environmental analysis. However, due to their low compatibility, it is difficult to use the common FABs for biological samples. To overcome this challenge, construction of FABs with complex structures to adapt the nature of biological samples is currently in progress in this field. Unlike previous works, we moved our range of vision from the FAB itself to the biological sample. Inspired by this idea, in this work, flat membrane-based liquid-phase microextraction (FM-LPME) with sufficient sample cleanup and preconcentration capacities was integrated with FABs. With the merits of both FM-LPME and FABs, the integrated LPME-FAB system displayed a clear synergistic enhancement for target analysis. Specifically, LPME in the LPME-FAB system provided purified and enriched Hg2+ for the FAB recognition, while the FAB recognition event promoted the extraction efficiency of LPME. Due to superior performances, the LPME-FAB system achieved highly sensitive analysis of Hg2+ in urine samples with a detection limit of 27 nM and accuracies in the range of 98-113%. To the best of our knowledge, this is the first time that an integrated LPME-FAB system was constructed for target analysis in biological samples. We believe that this study will provide a new insight into the next generation of biosensors, where the integration of sample preparation with detection probes is as important as the design of complex probes in the field of bioanalysis.

Removal of Polymerase Chain Reaction Inhibitors by Electromembrane Extraction.

Polymerase chain reaction (PCR) technology has become the cornerstone of DNA analysis. However, special samples (e.g., forensic samples, soil, food, and mineral medicine) may contain powerful PCR inhibitors. High levels of inhibitors can hardly be sufficiently removed by conventional DNA extraction approaches and may result in the complete failure of PCR. In this work, the removal of PCR inhibitors by electromembrane extraction (EME) was investigated for the first time. To demonstrate the universality of the approach, EME formats with and without supported membranes (termed parallel-EME and µ-EME, respectively) were employed, and both anionic [humic acid (HA)] and cationic (Ca2+) PCR inhibitors were used as models. During EME, charged inhibitors in the sample migrate into the liquid membrane in the presence of an electric field and might further leech into the waste solution, while PCR analytes remain in the sample. After EME, the clearance values for HA at 0.2 and 2.5 mg mL-1 were 94 and 85%, respectively, and that for Ca2+ (275 mM) was 63%. Forensic PCR-short tandem repeat (PCR-STR) genotyping showed that EME significantly reduced the interference by HA in PCR-STR analysis and displayed a higher HA purge capability compared to existing methods. Furthermore, by combining EME with liquid-liquid extraction or solid-phase extraction, satisfactory STR profiles were obtained from HA-rich blood samples. In addition, false-negative reports of bacterial detection in mineral medicine and shrimps were avoided after the removal of Ca2+ by µ-EME. Our research demonstrates the great potential of EME for the purification of DNA samples containing high-level PCR inhibitors and opens up a new application direction for EME.

Electromembrane Extraction and Mass Spectrometry for Liver Organoid Drug Metabolism Studies.

Liver organoids are emerging tools for precision drug development and toxicity screening. We demonstrate that electromembrane extraction (EME) based on electrophoresis across an oil membrane is suited for segregating selected organoid-derived drug metabolites prior to mass spectrometry (MS)-based measurements. EME allowed drugs and drug metabolites to be separated from cell medium components (albumin, etc.) that could interfere with subsequent measurements. Multiwell EME (parallel-EME) holding 100 µL solutions allowed for simple and repeatable monitoring of heroin phase I metabolism kinetics. Organoid parallel-EME extracts were compatible with ultrahigh-performance liquid chromatography (UHPLC) used to separate the analytes prior to detection. Taken together, liver organoids are well-matched with EME followed by MS-based measurements.

Green microfluidic liquid-phase microextraction of polar and non-polar acids from urine.

In this work, hippuric acid (log P = 0.5), anthranilic acid (log P = 1.3), ketoprofen (log P = 3.6), and naproxen (log P = 3.0) were simultaneously extracted by a green microfluidic device based on the principles of liquid-phase microextraction (LPME). Different deep eutectic solvents (DESs) were investigated as supported liquid membrane (SLM), and a mixture of camphor and menthol as eutectic solvents in the molar ratio 1:1 was found to be highly efficient for the simultaneous extraction of non-polar and polar acidic drugs. LPME was conducted for 6 min per sample. Urine sample was delivered to the system at 1 µL min-1, and target analytes were extracted exhaustively (75-100% recovery) across the DES SLM, and into pure aqueous phosphate buffer pH 11.0 delivered as acceptor at 1 µL min-1. The acceptor was analyzed with liquid chromatography-UV detection. Interestingly, the DES enabled extraction of both the polar and non-polar model analytes at the same time; all chemicals were green and non-hazardous, and the chemical waste was less than 1 mg per sample.

Selectivity and efficiency of electromembrane extraction of polar bases with different liquid membranes-Link to analyte properties.

In the present fundamental study, selectivity and efficiency of electromembrane extraction of 50 polar basic substances (-6.7 < log P < +1.0) was systematically studied for ten different supported liquid membranes. For each model substance, 23 molecular descriptors were collected and these were investigated as potential parameters for understanding of extraction efficiency and selectivity by means of partial least squares regression. Overall, a highly aromatic deep eutectic solvent composed of coumarin and thymol with addition of 2% ionic carrier (di(2-ethylhexyl) phosphate) provided the highest extraction efficiency with an average extraction yield of 69% from pure water samples, 55% from plasma, and 62% from urine. With this solvent system, ionic, cation-π, and π-π interactions between the supported liquid membrane and analytes were dominant. Supported liquid membranes without aromaticity, however, operated primarily based on hydrogen-bonding interactions. This is the first time the relationship between analyte properties, solvent composition, and extraction yield has systematically been studied for polar bases in electromembrane extraction. This new knowledge represents a first step toward enabling future development and optimization of electromembrane extraction systems for polar bases based on rational design, rather than trial-and-error approaches.

Electromembrane Extraction Using Sacrificial Electrodes.

In this paper, we report the first example of employing a sacrificial electrode in the acceptor solution during electromembrane extraction (EME). The electrode was based on a silver wire with a layer of silver chloride electroplated onto the surface. During EME, the electrode effectively inhibited electrolysis of water in the acceptor compartment, by accepting the charge transfer across the SLM, which enabled the application of 500 µA current without suffering gas formation or pH changes from electrolysis of water. The electroplating strategy was optimized with a design-of-experiments (DOE) methodology that provided optimal conditions of electroplating. With an optimized electrode, 1 cm of the electrode in contact with the acceptor solution inhibited electrolysis of water for approximately 30 min at 500 µA current (redox capacity). Further, the redox capacity of the electrode was found to increase through multiple uses. The advantage of the electrode was demonstrated by extracting polar analytes at high-current conditions in a standard EME system comprising 2-nitrophenyl octyl ether (NPOE) as SLM and 10 mM HCl as sample/acceptor solutions. Application of high current enabled significantly higher recoveries than could otherwise be obtained at 100 µA. Sacrificial electrodes were also tested in µ-EME and were found beneficial by eliminating detrimental bubble formation. Thus, the sacrificial electrodes improved the stability of µ-EME systems. The findings of this paper are important for development of stable and robust systems for EME operated at high voltage/current and for EME performed in narrow channels/tubing where bubble formation is critical.

3D cell culture models and organ-on-a-chip: Meet separation science and mass spectrometry.

In vitro derived simplified 3D representations of human organs or organ functionalities are predicted to play a major role in disease modeling, drug development, and personalized medicine, as they complement traditional cell line approaches and animal models. The cells for 3D organ representations may be derived from primary tissues, embryonic stem cells or induced pluripotent stem cells and come in a variety of formats from aggregates of individual or mixed cell types, self-organizing in vitro developed "organoids" and tissue mimicking chips. Microfluidic devices that allow long-term maintenance and combination with other tissues, cells or organoids are commonly referred to as "microphysiological" or "organ-on-a-chip" systems. Organ-on-a-chip technology allows a broad range of "on-chip" and "off-chip" analytical techniques, whereby "on-chip" techniques offer the possibility of real time tracking and analysis. In the rapidly expanding tool kit for real time analytical assays, mass spectrometry, combined with "on-chip" electrophoresis, and other separation approaches offer attractive emerging tools. In this review, we provide an overview of current 3D cell culture models, a compendium of current analytical strategies, and we make a case for new approaches for integrating separation science and mass spectrometry in this rapidly expanding research field.

Electromembrane extraction of sodium dodecyl sulfate from highly concentrated solutions.

This fundamental work investigated the removal of sodium dodecyl sulfate (SDS) from highly concentrated samples by electromembrane extraction (EME). SDS concentrations were in the range of 0.1-1.0% w/v, covering both sub- and super-critical micellar concentrations (CMC). Under optimal conditions, we extracted SDS from 100 µL aqueous sample, through 3 µL supported liquid membrane (SLM) and into 200 µL 10 mM NaOH in water as waste solution. The SLM comprised 1.0% w/w Aliquat 336 in 1-nonanol, and extraction voltage was 5 V. From 0.1% SDS samples, EME removed 100% during 30 minutes operation (100% clearance). SDS concentration above the critical micellar concentration (CMC) challenged the capacity of the system. Thus, to reach 100% clearance from 0.5% samples, we extracted for 120 minutes and replenished the SLM after 60 minutes. Increasing the membrane area of the SLM from 28 mm2 to 43 mm2 provided 100% clearance from 0.5% samples after 30 min EME. Complete clearance of 1.0% SDS samples was not achieved under the tested conditions, and maximal clearance was 60%. Mass balance experiments demonstrated that most of the removed SDS is trapped in the SLM, rather than transferring to the waste solution. For super-CMC samples, aggregation of SDS in the SLM exceeded the SLM capacity and impeded further mass transfer.

Influence of acid-base dissociation equilibria during electromembrane extraction.

Electromembrane extraction is affected by acid-base equilibria of the extracted substances as well as coupled equilibria associated with the partitioning of neutral substances to the supported liquid membrane. A theoretical model for this was developed and verified experimentally in the current work using pure 2-nitrophenyl octyl ether as supported liquid membrane. From this model, extraction efficiency as a function of pH can be predicted. Substances with log P < 0-2 are generally extracted with low efficiency. Substances with log P > 2 are generally extracted with high efficiency when acceptor pH < pKaH - log P. Twelve basic drug substances (2.07 < log P < 6.57 and 6.03 < pKaH  < 10.47) were extracted under different pH conditions with 2-nitrophenyl octyl ether as supported liquid membrane and fitted to the model. Seven of the drug substances behaved according to the model, while those with log P close to 2.0 deviated from prediction. The deviation was most probably caused by deprotonation and ion pairing within the supporting liquid membrane. Measured partition coefficients (log P) between 2-nitrophenyl octyl ether and water, were similar to traditional log P values between n-octanol and water. Thus, the latter have potential for pKaH - log P predictions.

Electromembrane Extraction of Unconjugated Fluorescein Isothiocyanate from Solutions of Labeled Proteins Prior to Flow Induced Dispersion Analysis.

In this initial research on feasibility, removal of unconjugated fluorescein isothiocyanate (FITC) after fluorescent labeling of human serum albumin (HSA) with electromembrane extraction (EME) was investigated for the first time. A 100 µL solution of 0.1 mg/mL HSA was fluorescently labeled with 0.01 mg/mL FITC in a molar ratio of 10:1 in an Eppendorf tube for 30 min under agitation and absence of light. Then the labeled solution was transferred to a 96-well EME with 3 µL 0.1% (w/w) Aliquat 336 in 1-octanol as the supported liquid membrane (SLM) and 200 µL 10 mM NaOH as waste solution. EME was performed for 10 min with a voltage of 50 V, with the anode in the waste solution and at 900 rpm agitation. Negatively charged and unconjugated FITC was extracted electrokinetically into the SLM and to the waste solution. Analysis of purified samples, by Taylor dispersion analysis (TDA), showed a 92% removal of unconjugated FITC (FITC clearance: 92%, RSD: 3%), while 79% of the HSA/FITC complex remained in the sample (protein retention: 79%, RSD: 18%). Conserved functionality of the HSA/FITC complex after EME was proven by a binding affinity study with anti-HSA using flow induced dispersion analysis (FIDA). In this real sample, the dissociation constant (Kd) and hydrodynamic radius of the complex were determined to be 0.8 µM and 5.87 nm, respectively, which was in concordance with previously reported values.

Liquid-Phase Microextraction or Electromembrane Extraction?

Isolation of substances by liquid-phase microextraction (LPME) or electromembrane extraction (EME) is becoming more and more important in analytical chemistry. However, the understanding of the mass transfer in LPME and EME is limited, especially for highly concentrated samples. In this work, the mass transfer in LPME and EME from aqueous samples (0.5-200 mg L-1) was studied in terms of recovery, equilibrium time, flux, and mass transfer capacity. In both EME and LPME, high recoveries were achieved at low analyte concentration, and the recoveries decreased at high analyte concentration. For EME, the loss in recovery was partly compensated by increasing the extraction voltage (from 50 to 200 V), while the LPME recovery at high analyte concentration was improved by increasing the extraction time (from 30 to 180 min). EME was superior in terms of equilibrium time and flux, while LPME provided much higher mass transfer capacity especially for highly concentrated samples. Moreover, the recovery was much more sensitive to high analyte concentrations in EME than in LPME, and the EME recovery decreased significantly above 50 mg L-1, indicating that LPME could be used to isolate analytes in a wider concentration range than EME. We believe that this fundamental study will be of great importance for the selection of a suitable membrane-based microextraction technique.

On-chip electromembrane extraction of acidic drugs.

In the present work, a new supported liquid membrane (SLM) has been developed for on-chip electromembrane extraction of acidic drugs combined with HPLC or CE, providing significantly higher stability than those reported up to date. The target analytes are five widely used non-steroidal anti-inflammatory drugs (NSAIDs): ibuprofen (IBU), diclofenac (DIC), naproxen (NAX), ketoprofen (KTP) and salicylic acid (SAL). Two different microchip devices were used, both consisted basically of two poly(methyl methacrylate) (PMMA) plates with individual channels for acceptor and sample solutions, respectively, and a 25 µm thick porous polypropylene membrane impregnated with the organic solvent in between. The SLM consisting of a mixture of 1-undecanol and 2-nitrophenyl octyl ether (NPOE) in a ratio 1:3 was found to be the most suitable liquid membrane for the extraction of these acidic drugs under dynamic conditions. It showed a long-term stability of at least 8 hours, a low system current around 20 µA, and recoveries over 94% for the target analytes. NPOE was included in the SLM to significantly decrease the extraction current compared to pure 1-undecanol, while the extraction properties was almost unaffected. Moreover, it has been successfully applied to the determination of the target analytes in human urine samples, providing high extraction efficiency.

Electromembrane extraction-looking into the future.

Analytical microextraction techniques, including solid-phase microextraction (SPME) Arthur & Pawliszyn (Anal Chem 62:2145-2148, 1990), stir bar sorptive extraction (SBSE), Baltussen et al. (J Microcol 11:737-747, 1999), single-drop microextraction (SDME) Jeannot & Cantwell (Anal Chem 68:2236-2240, 1996), hollow-fiber liquid-phase microextraction (HF-LPME) Pedersen-Bjergaard & Rasmussen (Anal Chem 71:2650-2656, 1999), dispersive liquid-liquid microextraction (DLLME) Berijani et al (J Chromatogr A. 1123:1-9, 2006), and electromembrane extraction (EME) Pedersen-Bjergaard & Rasmussen (J Chromatogr A 1109:183-190, 2006) have gained considerable interest in recent years. The latter technique, EME, differs from the others by the fact that mass transfer and extraction is facilitated by electrokinetic migration. Thus, basic or acidic analytes are extracted in their ionized form from aqueous sample, through an organic supported liquid membrane (SLM) and into an aqueous acceptor solution under the influence of an electrical potential. EME provides pre-concentration and sample clean-up, and can be performed in 96-well format using only a few microliter organic solvent per sample (green chemistry). Extraction selectivity is controlled by the direction and magnitude of the electrical field, by the chemical composition of the SLM, and by pH in the acceptor solution and sample. This trends article discusses briefly the principle, performance, and current status of EME, and from this future directions and perspectives are identified. Unlike traditional extraction methods, EME involves electrokinetic transfer of charged analyte molecules across an organic phase (SLM) immiscible with water. This process is still not fully characterized from a fundamental point of view, and more research in this area is expected in the near future. From author's point of view, such research at the interface between electrophoresis and partition will be highly important for future implementation of EME. Graphical abstract ᅟ.

Nanoliter-Scale Electromembrane Extraction and Enrichment in a Microfluidic Chip.

This paper reports for the first time nanoliter-scale electromembrane extraction (nanoliter-scale EME) in a microfluidic device. Six basic drug substances (model analytes) were extracted from 70 µL samples of human whole blood, plasma, or urine through a supported liquid membrane (SLM) of 2-nitrophenyl octyl ether (NPOE) and into 6 nL of 10 mM formic acid as an acceptor solution. A DC potential of 15 V was applied across the SLM and served as the driving force for the extraction. The cathode was located in the acceptor solution. Because of the small area of the SLM (0.06 mm2), the system provided soft extraction with recoveries <1% for the 70 µL samples. Because of the large sample-to-acceptor-volume ratio, analytes were enriched in the acceptor solution. The enrichment capacity was 6-7-fold per minute, and after 60 min of operation, most of the model analytes were enriched by a factor of approximately 400. Because of the SLM and the direction of the applied electrical field, substantial sample cleanup was obtained. The chips were based on thiol-ene polymers, and the soft-lithography-fabrication procedure and the materials were selected in such a way that future mass production should be feasible. The chip-to-chip variability was within 23% RSD (and less than 10% in most cases) with respect to extraction recovery. Our findings have verified that nanoliter-scale EME is highly feasible and provides reliable data, and for future studies, the concept should be tested for applicability in connection with in vitro microphysiological systems, organ-on-a-chip systems, and point-of-care diagnostics. These are potential areas where the combination of soft extraction and high enrichment from limited sample volumes is required for reliable analytical measurements.

Rapid determination of designer benzodiazepines, benzodiazepines, and Z-hypnotics in whole blood using parallel artificial liquid membrane extraction and UHPLC-MS/MS.

Benzodiazepines (BZD) and Z-hypnotics are frequently analyzed in forensic laboratories, and in 2012, the designer benzodiazepines (DBZD) emerged on the illegal drug scene. DBZD represent a particular challenge demanding new analytical methods. In this work, parallel artificial liquid membrane extraction (PALME) is used for sample preparation of DBZD, BZD, and Z-hypnotics in whole blood prior to UHPLC-MS/MS analysis. PALME of BZD, DBZD, and Z-hypnotics was performed from whole blood samples, and the analytes were extracted across a supported liquid membrane (SLM) and into an acceptor solution of dimethyl sulfoxide and 200 mM formic acid (75:25, v/v). The method was validated according to EMA guidelines. The method was linear throughout the calibration range (R2 > 0.99). Intra- and inter-day accuracy and precision, as well as matrix effects, were within the guideline limit of ± 15%. LOD and LLOQ ranged from 0.10 to 5.0 ng mL-1 and 3.2 to 160 ng mL-1, respectively. Extraction recoveries were reproducible and above 52%. The method was specific, and the analytes were stable in the PALME extracts for 4 and 10 days at 10 and - 20 °C. No carry-over was observed within the calibration range. PALME and UHPLC-MS/MS for the determination of DBZD, BZD, and Z-hypnotics in whole blood are a green and low-cost alternative that provides high sample throughput (96-well format), extensive sample clean-up, good sensitivity, and high reproducibility. The presented method is also the first method incorporating analysis of DBZD, BZD, and Z-hypnotics in whole blood in one efficient analysis. Graphical abstract.

Complexation-mediated electromembrane extraction of highly polar basic drugs-a fundamental study with catecholamines in urine as model system.

Complexation-mediated electromembrane extraction (EME) of highly polar basic drugs (log P < -1) was investigated for the first time with the catecholamines epinephrine, norepinephrine, and dopamine as model analytes. The model analytes were extracted as cationic species from urine samples (pH 4), through a supported liquid membrane (SLM) comprising 25 mM 4-(trifluoromethyl)phenylboronic acid (TFPBA) in bis(2-ethylhexyl) phosphite (DEHPi), and into 20 mM formic acid as acceptor solution. EME was performed for 15 min, and 50 V was used as extraction voltage across the SLM. TFPBA served as complexation reagent, and selectively formed boronate esters by reversible covalent binding with the model analytes at the sample/SLM interface. This enhanced the mass transfer of the highly polar model analytes across the SLM, and EME of basic drugs with log P in the range -1 to -2 was shown for the first time. Meanwhile, most matrix components in urine were unable to pass the SLM. Thus, the proposed concept provided highly efficient sample clean-up and the system current across the SLM was kept below 50 µA. Finally, the complexation-mediated EME concept was combined with ultra-high performance liquid chromatography coupled to tandem mass spectrometry and evaluated for quantification of epinephrine and dopamine. Standard addition calibration was applied to a pooled human urine sample. Calibration curves using standards between 25 and 125 µg L-1 gave a high level of linearity with a correlation coefficient of 0.990 for epinephrine and 0.996 for dopamine (N = 5). The limit of detection, calculated as three times signal-to-noise ratio, was 5.0 µg L-1 for epinephrine and 1.8 µg L-1 for dopamine. The repeatability of the method, expressed as coefficient of variation, was 13% (n = 5). The proposed method was finally applied for the analysis of spiked pooled human urine sample, obtaining relative recoveries of 91 and 117% for epinephrine and dopamine, respectively.