Three-phase Liquid-phase Microextraction Research Articles

Liquid-phase microextraction in bioanalysis – How green can it be?

• Liquid-phase microextraction of basic pharmaceuticals from plasma was optimized with respect to greenness • Traditional chemicals were replaced with green alternatives • Equipment was washed in hot water and recycled • Greenness was documented using the AGREEprep metric system Fundamental experiments were conducted with three-phase liquid-phase microextraction (LPME) to optimize greenness, followed by evaluation with the AGREEprep metric system. Extractions were from human plasma, and recoveries were studied for 90 different basic substances (drugs and endogenous metabolites) in the log P-range from -4.2 to 8.1. In a first set of experiments, LPME was performed in a 96-well system using traditional LPME chemicals. Extraction was from 125 µL of human plasma, diluted with 125 µL of 10 mM NaOH, through 3.0 µL of dihexyl ether as liquid membrane, and into 50 µL of 20 mM HCOOH as acceptor. The optimal extraction window (OEW) with this system ranged from log P 2.2 to 5.0, and the majority of drug substances within OEW were extracted exhaustively (average recovery 71 %). For this procedure, the AGREEprep metric score for greenness was 0.62. In a next set of experiments, NaOH, dihexyl ether, and HCOOH were replaced with NaHCO 3 , sesame oil, and citric acid to improve greenness. Even with sesame oil, the majority of the drug substances with log P from 2.2 to 5.0 were extracted with high efficiency, and the AGREEprep score increased to 0.68 due to the highly green nature of the chemicals. In a third set of experiments, where LPME was conducted in a vial-based approach, greenness was improved further by recycling and washing equipment in hot water, except the liquid membrane. Extraction performance was unaffected by washing and recycling, and no carry-over was observed from the extraction equipment. Upon equipment washing and recycling, the total material waste was reduced to less than 10 mg per sample, and the AGREEprep score increased to 0.72. Because experiments were conducted manually and in laboratory, and because LC-MS was used for final analysis, we were unable to improve the AGREEprep score further. The experiments illustrate that LPME procedures may be developed to a very high level of greenness, under assistance by the AGREEprep metric assessment approach.

Determination of tramadol hydrochloride in water samples by directly suspended droplet three-phase liquid phase microextraction with tetrathiocyanato cobalt(II) as the complexing agent using UV-spectroscopy

Suspended droplet three-phase liquid microextraction is a relatively new sample preparation technique for analysis of drugs. In the current study, tramadol hydrochloride was determined using tetrathiocyanato cobalt(II), (NH4)2[Co(SCN)4], as the complexing agent. The maximum of absorption for the formed ternary complex between tramadol hydrochloride and tetrathiocyanato cobalt(II) was observed at 625 nm. In this method, the target compound was extracted from the aqueous phase (5 mL containing 5 μg/mL of sample, pH 11) into an organic phase and then back-extracted into a directly suspended droplet as acceptor phase (3 μL of 1.74 × 10–9 M tetrathiocyanato cobalt(II), pH 2.8). 1-Octanol was used as organic phase. Under the optimized conditions: extraction time 90 s, back-extraction time 8 min and stirring rate 750 rpm, the linear range, the relative standard deviation (RSD, n = 5) and the limit of detection for tramadol hydrochloride were found 0.5–8 μg/mL, 4.9% and 8 ng/mL (n = 5), respectively. Also, the tap water and surface water samples were successfully analyzed using the proposed method and the relative recoveries obtained from the spiked water samples were 98.2 and 97.6%, respectively.

Hybrid flow analyzer for automatic hollow-fiber-assisted ionic liquid-based liquid-phase microextraction with in-line membrane regeneration

The proof-of-concept of a new methodology for in-line hollow-fiber (HF)-assisted three-phase liquid-phase microextraction (LPME) allowing for handling of the feed and acceptor aqueous solutions and of minute volumes of the organic extracting phase in a programmable flow mode is reported in this paper. The flow analyzer fosters in-line anchoring of ionic-liquid-laden extracting solution (10 % (v/v) methyltrioctyl ammonium chloride in kerosene) in the pores of a single-strand microporous polypropylene HF, and regeneration of the liquid-phase membrane itself for each individual analysis cycle in a fully automated mode. Using hexavalent chromium as a model analyte and 1,5-diphenylcarbazide as a chromogenic probe in the acceptor solution, the flow-based HF-LPME hyphenated system was harnessed to the clean-up of troublesome samples (viz., domestic wastewater and soil leachates) with concomitant enrichment of target species. Distinct extraction modes and chemistries were assessed for enhanced Cr(VI) permeability. A single sample plug was subjected to a twofold backward-forward flow extraction so as to decrease the thickness of the boundary layer at the HF shell side for improved extraction efficiency. Under the optimized physicochemical variables, a limit of detection of 4.6 μg L(-1) Cr(VI), a dynamic linear range of up to 500 μg L(-1) and intermediate precision better than 10 % were obtained for a sample volume of 2.8 mL buffered at pH 4 and a volume of organic extractant of 120 μL, with an enrichment factor of ca. 11 for a sample residence time in the donor compartment of merely 4.5 min. Analyte recoveries in domestic wastewaters were ≥83 % using external calibration with relative standard deviations better than 14 %, thereby demonstrating the expedient clean-up of samples with elevated content of dissolved organic carbon. The automatic HF-LPME method was validated in terms of bias against the SRM 2701 (NIST soil) preceded by the EPA alkaline digestion method 3060A. No significant differences between Cr(VI) concentration as obtained with the automatic HF-LPME system (546 ± 57 mg kg(-1)) and the certified value (551.2 ± 17.2 mg kg(-1), expressed as mean ± combined uncertainty) were encountered at the 0.05 significance level.

Liquid‐phase microextraction and desorption electrospray ionization mass spectrometry for identification and quantification of basic drugs in human urine

Hollow fibre liquid-phase microextraction (LPME) and desorption electrospray ionization mass spectrometry (DESI-MS) were evaluated for the identification and quantification of basic drugs in human urine samples. The selective extraction capabilities of three-phase LPME provided a significant reduction in the matrix effects otherwise observed in direct DESI-MS analysis of urine samples. Aqueous LPME extracts (in 10 mM HCl) were deposited on porous Teflon, dried at room temperature, and the dried spots were then analyzed directly with DESI-MS in full scan mode. Pethidine, diphenhydramine, nortriptyline, and methadone were used as model compounds for identification, and their limits of identification were determined to be 100, 25, 100, and 30 ng/mL, respectively. In a reliability test with 19 spiked urine samples, 100% of the positive samples containing the model drugs in concentrations at or above the limit of identification were identified. Diphenhydramine was used as a model compound for quantitative analysis with diphenhydramine-d(5) as an internal standard. The calibration curve was linear in the range 50-2000 ng/mL (R(2) = 0.992) with a limit of quantification at approximately 140 ng/mL. The intra- and inter-day relative standard deviations were <9.5%. In a reliability test with six spiked urine samples, deviations between the measured and the true values for diphenhydramine were in the range 0.2-22.9%.

Determination of partition coefficient and analysis of nitrophenols by three-phase liquid-phase microextraction coupled with capillary electrophoresis

A three-phase hollow fiber liquid-phase microextraction method coupled with CE was developed and used for the determination of partition coefficients and analysis of selected nitrophenols in water samples. The selected nitrophenols were extracted from 14 mL of aqueous solution (donor solution) with the pH adjusted to pH 3 into an organic phase (1-octanol) immobilized in the pores of the hollow fiber and finally backextracted into 40.0 microL of the acceptor phase (NaOH) at pH 12.0 located inside the lumen of the hollow fiber. The extractions were carried out under the following optimum conditions: donor solution, 0.05 M H(3)PO(4), pH 3.0; organic solvent, 1-octanol; acceptor solution, 40 microL of 0.1 M NaOH, pH 12.0; agitation rate, 1050 rpm; extraction time, 15 min. Under optimized conditions, the calibration curves for the analytes were linear in the range of 0.05-0.30 mg/L with r(2)>0.9900 and LODs were in the range of 0.01-0.04 mg/L with RSDs of 1.25-2.32%. Excellent enrichment factors of up to 398-folds were obtained. It was found that the partition coefficient (K(a/d)) values were high for 2-nitrophenol, 3-nitrophenol, 4-nitrophenol, 2,4-dinitrophenol and 2,6-dinitrophenol and that the individual partition coefficients (K(org/d) and K(a/org)) promoted efficient simultaneous extraction from the donor through the organic phase and further into the acceptor phase. The developed method was successfully applied for the analysis of water samples.

Hollow fiber-liquid-phase microextraction of fungicides from orange juices

Liquid-phase microextraction (LPME) based on polypropylene hollow fibers was evaluated for the extraction of the post-harvest fungicides thiabendazole (TBZ), carbendazim (CBZ) and imazalil (IMZ) from orange juices. Direct LPME was performed without any sample pretreatment prior to the extraction, using a simple home-built equipment. A volume of 500 μL of 840 mM NaOH was added to 3 mL of orange juice in order to compensate the acidity of the samples and to adjust pH into the alkaline region. Analytes were extracted in their neutral state through a supported liquid membrane (SLM) of 2-octanone into 20 μL of a stagnant aqueous solution of 10 mM HCl inside the lumen of the hollow fiber. Subsequently, the acceptor solution was directly subjected to analysis. Capillary electrophoresis (CE) was used during the optimization of the extraction procedure. Working under the optimized extraction conditions, LPME effectively extracted the analytes from different orange juices, regardless of different pH or solid material (pulp) present in the sample, with recoveries that ranged between 17.0 and 33.7%. The analytical performance of the method was evaluated by liquid chromatography coupled with mass spectrometry (LC/MS). This technique provided better sensitivity than CE and permitted the detection below the μg L −1 level. The relative standard deviations of the recoveries (RSDs) ranged between 3.4 and 10.6%, which are acceptable values for a manual microextraction technique without any previous sample treatment, using a home-built equipment and working under non-equilibrium conditions (30 min extraction). Linearity was obtained in the range 0.1–10.0 μg L −1, with r = 0.999 and 0.998 for TBZ and IMZ, respectively. Limits of detection were below 0.1 μg L −1 and are consistent with the maximum residue levels permitted for pesticides in drinking water, which is the most restrictive regulation applicable for these kinds of samples. It has been demonstrated the suitability of three-phase LPME for the extraction of pesticides from citrus juices, suppressing any pretreatment step such as filtration or removal of the solid material from the sample, that may potentially involve a loss of analyte.

Chiral HPLC analysis of venlafaxine metabolites in rat liver microsomal preparations after LPME extraction and application to an in vitro biotransformation study

A three-phase LPME (liquid-phase microextraction) method for the enantioselective analysis of venlafaxine (VF) metabolites (O-desmethylvenlafaxine (ODV) and N-desmethylvenlafaxine (NDV) in microsomal preparations is described for the first time. The assay involves the chiral HPLC separation of drug and metabolites using a Chiralpak AD column under normal-phase mode of elution and detection at 230 nm. The LPME procedure was optimized using multifactorial experiments and the following optimal condition was established: sample agitation at 1,750 rpm, 20 min of extraction, acetic acid 0.1 mol/L as acceptor phase, 1-octanol as organic phase and donor phase pH adjustment to 10.0. Under these conditions, the mean recoveries were 41% and 42% for (-)-(R)-ODV and (+)-(S)-ODV, respectively, and 47% and 48% for (-)-(R)-NDV and (+)-(S)-NDV, respectively. The method presented quantification limits of 200 ng/mL and it was linear over the concentration range of 200-5,000 ng/mL for all analytes. The validated method was employed to study the in vitro biotransformation of VF using rat liver microsomal fraction. The results demonstrated the enantioselective biotransformation of VF.

Determination of 3-nitroaniline in water samples by directly suspended droplet three-phase liquid-phase microextraction using 18-crown-6 ether and high-performance liquid chromatography

Liquid–liquid–liquid microextraction (LLLME) with directly suspended droplet in high-performance liquid chromatography (HPLC) has been applied as a new, rapid and easy method for the determination of 3-nitroaniline in environmental water samples. The target compound was extracted from the aqueous sample solution (donor phase, pH 13) into an organic phase and then was back-extracted into a directly suspended droplet of an acidic aqueous solution (acceptor phase, pH 0.3). In this method, without using a microsyringe as supporting device, an aqueous large droplet is freely suspended at the top-center position of an immiscible organic solvent, which is laid over the aqueous sample solution while being agitated. Then, the droplet was withdrawn into the microsyringe and directly was injected into the HPLC system with UV detection at 227 nm. Up to 148-fold enrichment of the analyte could be obtained under the optimal conditions [i.e. donor phase: 0.1 M sodium hydroxide solution (4.5 mL); organic phase: o-xylene/1-octanol (90:10, v/v; 250 μL); acceptor phase: 0.5 M hydrochloric acid and 500 mM 18-crown-6 ether (6 μL); extraction time: 60 s; back-extraction time: 6 min and stirring rate: 600 rpm]. The limit of detection was 1 μg/L ( n = 7) and the relative standard deviation (RSD, n = 5) was 4.9 at S/N = 3. The calibration graph was linear in the range of 5–1500 μg/L with r = 0.9983. All experiments were carried out at room temperature (22 ± 0.5 °C).

Enantioselective determination of chloroquine and its n‐dealkylated metabolites in plasma using liquid‐phase microextraction and LC‐MS

A selective method using three-phase liquid-phase microextraction (LPME) in conjunction with LC-MS-MS was devised for the enantioselective determination of chloroquine and its n-dealkylated metabolites in plasma samples. After alkalinization of the samples, the analytes were extracted into n-octanol immobilized in the pores of a polypropylene hollow fiber membrane and back extracted into the acidic acceptor phase (0.1 M TFA) filled into the lumen of the hollow fiber. Following LPME, the analytes were resolved on a Chirobiotic V column using methanol/ACN/glacial acetic acid/diethylamine (90:10:0.5:0.5 by volume) as the mobile phase. The MS detection was carried out using multiple reaction monitoring with ESI in the positive ion mode. The optimized LPME method yielded extraction recoveries ranging from 28 to 66%. The method was linear over 5-500 ng/mL and precision (RSD) and accuracy (relative error) values were below 15% for all analytes. The developed method was applied to the determination of the analytes in rat plasma samples after oral administration of the racemic drug.

Enantioselective analysis of mirtazapine and its two major metabolites in human plasma by liquid chromatography–mass spectrometry after three-phase liquid-phase microextraction

A three-phase liquid-phase microextraction (LPME) method using porous polypropylene hollow fibre membrane with a sealed end was developed for the extraction of mirtazapine (MRT) and its two major metabolites, 8-hydroxymirtazapine (8-OHM) and demethylmirtazapine (DMR), from human plasma. The analytes were extracted from 1.0 mL of plasma, previously diluted and alkalinized with 3.0 mL 0.5 mol L −1 pH 8 phosphate buffer solution and supplemented with 15% sodium chloride (NaCl), using n-hexyl ether as organic solvent and 0.01 moL L −1 acetic acid solution as the acceptor phase. Haloperidol was used as internal standard. The chromatographic analyses were carried out on a chiral column, using acetonitrile–methanol–ethanol (98:1:1, v/v/v) plus 0.2% diethylamine as mobile phase, at a flow rate of 1.0 mL min −1. Multi-reaction monitoring (MRM) detection was performed by mass spectrometry (MS–MS) using a triple-stage quadrupole and electrospray ionization interface operating in the positive ion mode. The mean recoveries were in 18.3–45.5% range with linear responses over the 1.25–125 ng mL −1 concentration range for all enantiomers evaluated. The quantification limit (LOQ) was 1.25 ng mL −1. Within-day and between-day assay precision and accuracy (2.5, 50 and 100 ng mL −1) showed relative standard deviation and the relative error lower than 11.9% for all enantiomers evaluated. Finally, the method was successfully used for the determination of mirtazapine and its metabolite enantiomers in plasma samples obtained after single drug administration of mirtazapine to a healthy volunteer.

Supported liquid membranes in hollow fiber liquid‐phase microextraction (LPME) – Practical considerations in the three‐phase mode

In this work, three-phase liquid-phase microextraction (LPME) based on a supported liquid membrane (SLM) sustained in the wall of a hollow fiber was investigated with special focus on optimization of the experimental procedures in terms of recovery and repeatability. Recovery data for doxepin, amitriptyline, clomipramine, and mianserin were in the range of 67.8-79.8%. Within-day repeatability data for the four basic drugs were in the range of 4.1-7.7%. No single factor was found to be responsible for these variations, and the variability was caused by several factors related to the LPME extractions as well as to the final HPLC determination. Although the volume of the SLM varied within 0.4-3.1% RSD depending on the preparation procedure, and the volume of the acceptor solution varied within 4.8% RSD, both recoveries and repeatability were found to be relative insensitive to these variations. Thus, the handling of microliters of liquid in LPME was not a very critical factor, and the preparation of the SLM was accomplished in several different ways with comparable performance. Reuse of hollow fibers was found to suffer from matrix effects due to built-up of analytes in the SLM, whereas washing of the hollow fibers in acetone was beneficial in terms of recovery, especially for the extraction of the most hydrophobic substances. Several of the organic solvents used in the literature as SLM suffered from poor long-term stability, but silicone oil AR 20 (polyphenylmethylsiloxane), 2-nitrophenyl octyl ether (NPOE), and dodecyl acetate (DDA) all extracted with unaltered performance even after 60 days of storage at room temperature.

Liquid-phase microextraction combined with high-performance liquid chromatography for the enantioselective analysis of mefloquine in plasma samples

A simple and rapid method, which involves liquid-phase microextraction (LPME) followed by HPLC analysis using Chiralpak AD column and UV detection, was developed for the enantioselective determination of mefloquine in plasma samples. Several factors that influence the efficiency of three-phase LPME were investigated and optimized. Under the optimal extraction conditions, the mean recoveries were 33.2 and 35.0% for (−)-( SR-)-mefloquine and (+)-( RS)-mefloquine, respectively. The method was linear over 50–1500 ng/ml range. Within-day and between-day assay precision and accuracy were below 15% for both enantiomers at concentrations of 150, 600 and 1200 ng/ml. Furthermore, no racemization or degradation were seen with the method described.

Antibiotic-assisted three-phase liquid-phase microextraction of aromatic amines from aqueous solutions combined with high-performance liquid chromatography

Three-phase liquid-phase microextraction (LLLME) combined with high-performance liquid chromatography (HPLC) equipped with a monolithic column for the analysis of some aromatic amines is described. These compounds were extracted from an aqueous sample (4.0 mL) adjusted to pH 13 with NaOH-NaCl solution (Donor Phase, P1) into an organic phase (P2, 150 μL of Benzyl alcohol and ethyl acetate mixture, 2:1) and then back-extracted into a microdrop of aqueous acceptor phase (P3) adjusted to pH 2 with Na2HPO4-H3PO4 buffer solution. The extraction time T1 (from P1 to P2) was 20 min, and T2 (from P2 to P3) was 1 min. Different antibiotics as complexing agents for amines were added to the acceptor phase to improve the extraction time. Factors such as organic solvents, extraction times, addition of antibiotics to the acceptor phase, and stirring rate were optimized. The method was applied to the determination of aromatic amines in waste-water samples. Enrichment factors ranged from 189.0 to 395.5. The linearity range was from 2.5 to 1000 ng/mL, and the detection limits varied from 0.5 to 1.50 ng/mL. Relative standard deviations (%, n = 5) were found (at S/N 3) to be in the range of 1.3–8.5. All experiments were carried out at room temperature, 22 ± 0.5°C.

Experiences with Carrier-Mediated Transport in Liquid-Phase Microextraction

Different organic borates, phosphates, sulphates, and carboxylic acids are evaluated as extraction carriers in three-phase liquid-phase microextraction (LPME). Hydrophilic basic drugs form ion-pairs with the carriers and are extracted as ion-pair complexes into an organic liquid membrane of n-octanol or peppermint oil immobilized in the pores of a polypropylene hollow fiber. From this point, the basic drugs are released into a 20-microL solution of 50mM HCl placed inside the lumen of the hollow fiber (acceptor solution). Simultaneously, the carrier is neutralized by protons from the acceptor solution (protonated to maintain the charge balance). Both water-soluble and water-insoluble carriers are tested. One promising candidate among the water-soluble carriers is 1-heptanesulfonic acid. This is added to the sample solution to a final concentration of 25mM and served to ion-pair the analytes within the sample solution. Among the less water-soluble candidates, a mixture of di(2-ethylhexyl) phosphate (DEHP) and tris(2-ethylhexyl) phosphate (TEHP) serve as efficient carriers. Ten percent (w/w) of each of DEHP and TEHP are added to the organic liquid membrane, and these carriers principally worked through ion-pairing with the analytes at the interface between the sample solution and the organic liquid membrane. Several carriers are found to be compatible with human plasma samples, and bromthymol blue is particularly efficient in combination with these protein-containing matrices. Following optimization of the conditions for bromthymol blue, including saturation of the plasma samples with sodium sulphate, extraction recoveries between 45% and 75% are obtained for eight model drugs after 60 min of extraction. With bromthymol blue as the carrier, highly acceptable validation data are obtained for phenylpropanolamine and practolol extracted from human plasma.

Extraction and preconcentration of salbutamol and terbutaline from aqueous samples using hollow fiber supported liquid membrane containing anionic carrier

This paper presents a new three-phase liquid-phase microextraction (LPME) strategy for extraction and preconcentration of salbutamol (SB) and terbutaline (TB) from aqueous samples, including urine. The drugs were extracted from 11 ml of aqueous sample (source phase; SP) into an organic phase with microliter volume located inside the pores of a polypropylene hollow fiber, and then back-extracted into 24 μl of a second aqueous solution as the receiving phase (RP), located in the lumen of the hollow fiber. In preliminary experiments, we tried to transport the drugs using a pH gradient between the two sides of the hollow fiber. Due to the existence of both amine and phenolic groups on the drugs, very little transport occurred and enrichment factors (EF) less than one were obtained. Further experiments were done in the presence of bis(2-ethylhexyl) monohydrogenphosphoric acid (D2EHPA) or methyltrioctylammonium chloride (Aliquat 336) in the organic phase, to extract drugs from acidic and basic matrices, respectively. Results showed that transport of drugs from alkaline solution into 1 M of sodium bromide occurred when the membrane was impregnated with dihexyl ether containing 20% Aliquat 336. To optimize the EF, the effects of different parameters such as the nature of organic solvent used to impregnate the membrane, compositions and volumes of SP and RP, type and concentration of carrier, extraction time and stirring rate were investigated. Optimal results were obtained in the presence of 0.005 M of NaOH (pH 11.70) in the SP, 1 M of NaBr in the RP, 20% of Aliquat 336 in dihexyl ether as membrane impregnation solvent, stirring rate of 500 rpm and extraction time of 60 min. Under these conditions, enrichment factors of 52.9 and 213.1, dynamic linear ranges of 20–5000 and 10–5000, and limits of detection of 2.5 and 0.5 ng/ml were obtained for salbutamol and terbutaline, respectively. Also determination of drugs in environmental water and urine samples in the range of nanograms per millilitre with RSDs < 10% was possible using HPLC–photodiode array detection or HPLC–MS.

Ion-pair mediated transport of angiotensin, neurotensin, and their metabolites in liquid phase microextraction under acidic conditions

This paper discusses the behaviour of angiotensin 1 and neurotensin together with their metabolites in a three-phase liquid phase microextraction under acidic conditions. Variations in donor phase, organic phase, and acceptor phase are studied with extraction recovery as response variable. It is proved that for all peptides the transport across the organic phase is mediated by heptane-1-sulphonic acid. n-Octanol gave overall best results as organic phase. A donor phase volume of 1.0 mL was chosen as a compromise between optimal recovery and robustness of the LPME device. The optimal pH of the donor phase (using acceptor phase of pH 2) was found to be different for the peptides, which opens opportunities for selective sample preparation. Decreasing the acceptor phase pH to 1.0 resulted in increased extraction recoveries. On using 1.0 mL of donor phase containing 50 mM heptane-1-sulphonic acid pH 3, n-octanol as organic phase immobilized in the pores of the fibre, and 20 microL of acceptor phase containing 0.1 mol/L HCl, extraction recoveries up to 82% (enrichment factor = 41) were achieved. To our knowledge this is the first report on liquid phase microextraction of angiotensins and neurotensins.

Three-phase liquid-phase microextraction of weakly basic drugs from whole blood

The behaviour of weak basic analytes in liquid-phase microextraction (LPME) and the optimisation of parameters in whole blood are described. Benzodiazepines and non-benzodiazepine drugs were chosen as model substances. Liquid-phase microextraction based on disposable polypropylene hollow fibres was used in the three-phase extraction of five weak bases from whole blood. The sample work up with the liquid-phase microextraction technique can be impeded by low recovery due to incomplete trapping in the acceptor phase of weakly basic drugs and the complexity of the whole blood matrix. Different parameters related to this problem were experimentally studied. Additionally the stability of the analytes was examined because of low pH in the acceptor phase. The investigation resulted in optimised LPME conditions for the extraction of weak bases from whole blood. The parameters limiting the recovery were evaluated.

Recovery, enrichment and selectivity in liquid-phase microextraction: Comparison with conventional liquid–liquid extraction

Mathematical descriptions for extraction recovery and enrichment were applied for liquid-phase microextraction (LPME) and comparison with conventional two- and three-phase liquid–liquid extraction techniques (LLE) was made. The LPME theoretical calculations were verified by experimental determination of actual partition coefficients and by data obtained with LPME in a robust hollow fibre formate. With hollow fibre LPME operated in the two-phase mode, analytes were extracted from 1 to 4 ml aqueous samples into 25–50 μl of an organic solvent present in the pores and in the lumen of the porous hollow fibres. Compared with conventional two-phase LLE, two-phase LPME provided substantially higher enrichments for compounds with relatively large partition coefficients ( K org/d>500). In contrast, because of the large volume of organic solvent relative to the sample volume, LLE provided high recovery and moderate enrichment even for compounds with relatively low partition coefficients ( K org/d>5). Thus, two-phase LPME may be used for substantially enhanced extraction selectivity and enrichment of relatively hydrophobic analytes as compared with LLE whereas conventional two-phase LLE is superior for more hydrophilic analytes. Similar results were found for three-phase LPME where analytes where extracted from 1 to 4 ml aqueous samples through approximately 20 μl organic solvent immobilized within the pores of the hollow fibre and into 25 μl of an aqueous acceptor solution inside the lumen of the hollow fibre. The fundamental differences of LPME and LLE were further demonstrated with practical experiments on extraction of the basic drugs promethazine, methadone, and haloperidol from human plasma and urine.