Evolution of dispersive liquid–liquid microextraction method

Dispersive liquid–liquid microextraction (DLLME) has become a very popular environmentally benign sample-preparation technique, because it is fast, inexpensive, easy to operate with a high enrichment factor and consumes low volume of organic solvent. DLLME is a modified solvent extraction method in which acceptor-to-donor phase ratio is greatly reduced compared with other methods. In this review, in order to encourage further development of DLLME, its combination with different analytical techniques such as gas chromatography (GC), high-performance liquid chromatography (HPLC), inductively coupled plasma-optical emission spectrometry (ICP-OES) and electrothermal atomic absorption spectrometry (ET AAS) will be discussed. Also, its applications in conjunction with different extraction techniques such as solid-phase extraction (SPE), solidification of floating organic drop (SFO) and supercritical fluid extraction (SFE) are summarized. This review focuses on the extra steps in sample preparation for application of DLLME in different matrixes such as food, biological fluids and solid samples. Further, the recent developments in DLLME are presented. DLLME does have some limitations, which will also be discussed in detail. Finally, an outlook on the future of the technique will be given.     

Application of dispersive liquid–liquid microextraction with alcoholic solvents followed by HPLC–UV as a sensitive and efficient method for the extraction and determination of citalopram in biological samples using an experimental design

A sensitive and rapid method based on alcoholic-assisted dispersive liquid–liquid microextraction followed by high-performance liquid chromatography for determination of citalopram in human plasma and urine samples was developed. The effects of six parameters (extraction time, stirring speed, pH, volume of extraction and disperser solvents, and ionic strength) on the extraction recovery were investigated and optimized utilizing Plackett–Burman design and Box–Behnken design, respectively. According to Plackett–Burman design results, the volume of disperser solvent, stirring speed, and extraction time had no effect on the recovery of citalopram. The optimized condition was a mixture of 172 µL of 1-octanol as extraction solvent and 400 µL of methanol as disperser solvent, pH of 10.3 and 1% w/v of salt in the sample solution. Replicating the experiment in optimized condition for five times, gave the average extraction recoveries equal to 89.42%. The detection limit of citalopram in human plasma was obtained 4 ng/mL, and the linearity was in the range of 10–1200 ng/mL. The corresponding values for human urine were 5.4 ng/mL with the linearity in the range of 10–2000 ng/mL. Relative standard deviations for inter- and intraday extraction of citalopram were less than 7% for five measurements. The proposed method was successfully implemented for the determination of citalopram in human plasma and urine samples.     

Development of a dispersive liquid–liquid microextraction method for the determination of fluoroquinolones in chicken liver by high performance liquid chromatography

A simple and cost effective sample pre-treatment method, dispersive liquid–liquid microextraction (DLLME), has been developed for the extraction of six fluoroquinolones (FQs) from chicken liver samples. Clean DLLME extracts were analyzed for fluoroquinolones using liquid chromatography with diode array detection (LC-DAD). Parameters such as type and volume of disperser solvent, type and volume of extraction solvent, concentration and composition of phosphoric acid in the disperser solvent and pH were optimized. Linearity in the concentration range of 30–500 μg kg⁻¹ was obtained with regression coefficients ranging from 0.9945 to 0.9974. Intra-day repeatability expressed as % RSD was between 4 and 7%. The recoveries determined in spiked blank chicken livers at three concentration levels (i.e. 50, 100 and 300 μg kg⁻¹) ranged from 83 to 102%. LODs were between 5 and 19 μg kg⁻¹ while LOQs ranged between 23 and 62 μg kg⁻¹. All of the eight chicken liver samples obtained from the local supermarkets were found to contain at least one type of fluoroquinolone with enrofloxacin being the most commonly detected. Only one sample had four fluoroquinolone antibiotics (ciprofloxacin, difloxacin, enrofloxacin, norfloxacin). Norfloxacin which is unlicensed for use in South Africa was also detected in three of the eight chicken liver samples analyzed. The concentration levels of all FQs antibiotics in eight samples ranged from 8.8 to 35.3 μg kg⁻¹, values which are lower than the South African stipulated maximum residue limits (MRL).     

Solid-phase extraction combined with dispersive liquid–liquid microextraction for the determination for polybrominated diphenyl ethers in different environmental matrices

An analytical method, solid-phase extraction combined with dispersive liquid–liquid microextration (SPE–DLLME), was established to determine polybrominated diphenyl ethers (PBDEs) in water and plant samples. After concentration and purification of the samples in LC-C18 column, 1.0-mL elution sample containing 22.0 μL 1,1,2,2-tetrachloroethane was injected rapidly into the 5.0-mL pure water. After extraction and centrifuging, the sedimented phase was injected rapidly into gas chromatography with electron-capture detection (GC–ECD). For water samples, enrichment factors (EFs) are in the range of 6838–9405 under the optimum conditions. The calibration curves are linear in the range of 0.1–100 ng L⁻¹ (BDEs 28, 47) and 0.5–500 ng L⁻¹ (BDEs 100, 99, 85, 154, 153). The relative standard deviations (RSDs) and the limits of detection (LODs) are in the range of 4.2–7.9% (n = 5) and 0.03–0.15 ng L⁻¹, respectively. For plant samples, RSDs and LODs are in the range of 5.9–11.3% and 0.04–0.16 μg kg⁻¹, respectively. The relative recoveries of well, river, sea, leachate, and clover samples, spiked with different levels of PBDEs, are 66.8–94.1%, 72.2–100.5%, 74.5–110.4%, 62.1–105.1%, 66.1–91.7%, 62.4–88.9%, and 64.5–83.2%, respectively. The results show that SPE–DLLME is a suitable method for the determination of PBDEs in water and plant samples.     

Homogeneous liquid–liquid microextraction via flotation assistance for rapid and efficient determination of polycyclic aromatic hydrocarbons in water samples

In this work, a rapid, simple and efficient homogeneous liquid–liquid microextraction via flotation assistance (HLLME-FA) method was developed based on applying low density organic solvents without no centrifugation. For the first time, a special extraction cell was designed to facilitate collection of the low-density solvent extraction in the determination of four polycyclic aromatic hydrocarbons (PAHs) in water samples followed by gas chromatography-flame ionization detector (GC-FID). The effect of different variables on the extraction efficiency was studied simultaneously using experimental design. The variables of interest in the HLLME-FA were selected as extraction and homogeneous solvent volumes, ionic strength and extraction time. Response surface methodology (RSM) was applied to investigate the optimum conditions of all the variables. Using optimized variables in the extraction process, for all target PAHs, the detection limits, the precisions and the linearity of the method were found in the range of 14–41 μg L⁻¹, 3.7–10.3% (RSD, n = 3) and 50–1000 μg L⁻¹, respectively. The proposed method has been successfully applied to the analysis of four target PAHs in the water samples, and satisfactory results were obtained.     

Ultrasound-assisted dispersive liquid–liquid microextraction for the determination of six pyrethroids in river water

A simple ultrasound-assisted dispersive liquid–liquid microextraction method combined with liquid chromatography was developed for the preconcentration and determination of six pyrethroids in river water samples. The procedure was based on a ternary solvent system to formatting tiny droplets of extractant in sample solution by dissolving appropriate amounts of water-immiscible extractant (tetrachloromethane) in watermiscible dispersive solvent (acetone). Various parameters that affected the extraction efficiency (such as type and volume of extraction and dispersive solvent, extraction time, ultrasonic time, and centrifuging time) were evaluated. Under the optimum condition, good linearity was obtained in a range of 0.00059–1.52 mg L⁻¹ for all analytes with the correlation coefficient (r²) > 0.999. Intra-assay and inter-assay precision evaluated as the relative standard deviation (RSD) were less than 3.4 and 8.9%. The recoveries of six pyrethroids at three spiked levels were in the range of 86.2–109.3% with RSD of less than 8.7%. The enrichment factors for the six pyrethroids were ranged from 767 to 1033 folds.     

Development of a dispersive liquid–liquid microextraction method for determination of palladium in water samples using dicyclohexano-18- crown-6 as extracting agent

A new separation procedure for determination of palladium using dispersive liquid–liquid microextraction with dicyclohexano-18-crown-6 as complexing reagent was developed. In this method, potassium–dicyclohexano-18-crown-6 was used as a hydrophobic complex for the microextraction of palladium as PdCl₄ ^2? complex ion. The main factors affecting DLLME efficiency, such as type and volume of extractant and disperser solvent, concentration of chelating reagent, concentration of KCl and pH were optimized. Under the optimal conditions, the limit of detection for palladium was 16.0 ng mL^?1 with enrichment factor of 138. The present method was applied to the determination of palladium in water samples with satisfactory analytical results. The method was simple, rapid, cost efficient and sensitive for the extraction and preconcentration of palladium.     

Dispersive liquid–liquid microextraction coupled to liquid chromatography for thiamine determination in foods

A miniaturized dispersive liquid–liquid microextraction (DLLME) procedure coupled to liquid chromatography (LC) with fluorimetric detection was evaluated for the preconcentration and determination of thiamine (vitamin B₁). Derivatization was carried out by chemical oxidation of thiamine with 5 × 10⁻⁵ M ferricyanide at pH 13 to form fluorescent thiochrome. For DLLME, 0.5 mL of acetonitrile (dispersing solvent) containing 90 μL of tetrachloroethane (extraction solvent) was rapidly injected into 10 mL of sample solution containing the derivatized thiochrome and 24% (w/v) sodium chloride, thereby forming a cloudy solution. Phase separation was carried out by centrifugation, and a volume of 20 μL of the sedimented phase was submitted to LC. The mobile phase was a mixture of a 90% (v/v) 10 mM KH₂PO₄ (pH 7) solution and 10% (v/v) acetonitrile at 1 mL min⁻¹. An amide-based stationary phase involving a ligand with amide groups and the endcapping of trimethylsilyl was used. Specificity, linearity, precision, recovery, and sensitivity were satisfactory. Calibration graph was carried out by the standard additions method and was linear between 1 and 10 ng mL⁻¹. The detection limit was 0.09 ng mL⁻¹. The selectivity of the method was judged from the absence of interfering peaks at the thiamine elution time for blank chromatograms of unspiked samples. A relative standard deviation of 3.2% was obtained for a standard solution containing thiamine at 5 ng mL⁻¹. The esters thiamine monophosphate and thiamine pyrophosphate can also be determined by submitting the sample to successive acid and enzymatic treatments. The method was applied to the determination of thiamine in different foods such as beer, brewer’s yeast, honey, and baby foods including infant formulas, fermented milk, cereals, and purees. For the analysis of solid samples, a previous extraction step was applied based on an acid hydrolysis with trichloroacetic acid. The reliability of the procedure was checked by analyzing a certified reference material, pig’s liver (CRM 487). The value obtained was 8.76 ± 0.2 μg g⁻¹ thiamine, which is in excellent agreement with the certified value, 8.6 ± 1.1 μg g⁻¹.     

Coupling of air-assisted liquid–liquid microextraction method with partial least squares for simultaneous spectrophotometric determination of some preservatives

An air-assisted liquid–liquid microextraction method coupled with a multivariate calibration method, namely partial least squares (PLS), was developed for the extraction and simultaneous determination of benzoic acid (BA) and sorbic acid (SA) via a spectrophotometric approach. In this work, a two-step microextraction method was used. In the first step, analytes were extracted from acidic aqueous solution into octanol, as an organic solvent, and in the second step, the analytes were simultaneously back-extracted into an alkaline aqueous solution. The high absorption signal of octanol was the main reason to perform this back-extraction step. The effects of different parameters on the method efficiency were investigated; the parameters included extraction solvent volume, ionic strength of aqueous solution, pH, number of extraction cycles, and aqueous sample volume. Under optimum conditions, calibration graphs were seen to be linear over the range of 0.1–2.0 µg mL^?1 for the both analytes. Other analytical parameters were obtained as follows: Enrichment factors (EFs) were found to be 14.98 and 13.03, and limits of detection were determined to be 0.03 and 0.04 µg mL^?1 for BA and SA, respectively. As the last step, binary mixtures of the analytes were prepared and simultaneously extracted using the proposed method. Finally, PLS modeling was used for multivariate calibration of spectrophotometric data. It was successfully utilized for the analysis of the target analytes in real samples.     

Development of an ionic liquid based dispersive liquid–liquid microextraction method for the analysis of polycyclic aromatic hydrocarbons in water samples

A simple, rapid and efficient method, ionic liquid based dispersive liquid–liquid microextraction (IL-DLLME), has been developed for the first time for the determination of 18 polycyclic aromatic hydrocarbons (PAHs) in water samples. The chemical affinity between the ionic liquid (1-octyl-3-methylimidazolium hexafluorophosphate) and the analytes permits the extraction of the PAHs from the sample matrix also allowing their preconcentration. Thus, this technique combines extraction and concentration of the analytes into one step and avoids using toxic chlorinated solvents. The factors affecting the extraction efficiency, such as the type and volume of ionic liquid, type and volume of disperser solvent, extraction time, dispersion stage, centrifuging time and ionic strength, were optimised. Analysis of extracts was performed by high performance liquid chromatography (HPLC) coupled with fluorescence detection (Flu). The optimised method exhibited a good precision level with relative standard deviation values between 1.2% and 5.7%. Quantification limits obtained for all of these considered compounds (between 0.1 and 7 ng L⁻¹) were well below the limits recommended in the EU. The extraction yields for the different compounds obtained by IL-DLLME, ranged from 90.3% to 103.8%. Furthermore, high enrichment factors (301–346) were also achieved. The extraction efficiency of the optimised method is compared with that achieved by liquid–liquid extraction. Finally, the proposed method was successfully applied to the analysis of PAHs in real water samples (tap, bottled, fountain, well, river, rainwater, treated and raw wastewater).     

The recent developments in dispersive liquid–liquid microextraction for preconcentration and determination of inorganic analytes

Recently, increasing interest on the use of dispersive liquid–liquid microextraction (DLLME) developed in 2006 by Rezaee has been found in the field of separation science. DLLME is miniaturized format of liquid–liquid extraction in which acceptor-to-donor phase ratio is greatly reduced compared with other methods. In the present review, the combination of DLLME with different analytical techniques such as atomic absorption spectrometry (AAS), inductively coupled plasma-optical emission spectrometry (ICP-OES), gas chromatography (GC), and high-performance liquid chromatography (HPLC) for preconcentration and determination of inorganic analytes in different types of samples will be discussed. Recent developments in DLLME, e.g., displacement-DLLME, the use of an auxiliary solvent for adjustment of density of extraction mixture, and the application of ionic liquid-based DLLME in determination of inorganic species even in the presence of high content of salts are presented in the present review. Finally, comparison of DLLME with the other liquid-phase microextraction approaches and limitations of this technique are provided.     

New temperature-assisted ionic liquid-based dispersive liquid–liquid microextraction method for the determination of glyphosate and aminomethylphosphonic acid in water samples

A new analytical temperature-assisted ionic liquid-based dispersive liquid–liquid microextraction (TA-IL-DLLME) method was developed for glyphosate and aminomethylphosphonic acid determination in water samples. Extracted analytes were derivatized using 9-fluoroenylmethylchloroformate and quantified by liquid chromatography with fluorescence detection. For the TA-IL-DLLME method, two strategies for phase solubilization were evaluated; in approach 1, the ionic liquid and aqueous matrix sample were mixed and then heated, while in approach 2, the aqueous sample was first heated and then the ionic liquid was injected. For both approaches, optimization included parameters that significantly affect extraction efficiency: ionic liquid type and volume, solubilization temperature and time, cooling and centrifugation time. Among the evaluated ionic liquids, 1-decyl-3-methylimidazolium tetrafluoroborate showed the best performance for TA-IL-DLLME and was selected for the two solubilization approaches; with approach 2, slightly better results were obtained. Thus, sample analyses were performed using a procedure based on approach 2. An important matrix effect, attributed to the presence of salts and metals in real water samples was observed. Sample acidification before derivatization allowed this problem to diminish, with recoveries ranging from 75 and 99%, and enrichment factors between 57 and 76 for target analytes.     

Dispersive liquid–liquid microextraction and spectrophotometric determination of cobalt in water samples

A new simple and rapid dispersive liquid–liquid microextraction has been applied to preconcentrate trace levels of cobalt as a prior step to its determination by spectrophotometric detection. In this method a small amount of chloroform as the extraction solvent was dissolved in pure ethanol as the disperser solvent, then the binary solution was rapidly injected by a syringe into the water sample containing cobalt ions complexed by 1-(2-pyridylazo)-2-naphthol (PAN). This forms a cloudy solution. The cloudy state was the result of chloroform fine droplets formation, which has been dispersed in bulk aqueous sample. Therefore, Co-PAN complex was extracted into the fine chloroform droplets. After centrifugation (2 min at 5000 rpm) these droplets were sedimented at the bottom of conical test tube (about 100 µL) and then the whole of complex enriched extracted phase was determined by a spectrophotometer at 577 nm. Complex formation and extraction are usually affected by some parameters, such as the types and volumes of extraction solvent and disperser solvent, salt effect, pH and the concentration of chelating agent, which have been optimised for the presented method. Under optimum conditions, the enhancement factor (as the ratio of slope of preconcentrated sample to that obtained without preconcentration) of 125 was obtained from 50 mL of water sample, and the limit of detection (LOD) of the method was 0.5 µg L^?1and the relative standard deviation (RSD, n = 5) for 50 µg L^?1 of cobalt was 2.5%. The method was applied to the determination of cobalt in tap and river water samples.     

Dispersive liquid–liquid–liquid microextraction combined with liquid chromatography for the determination of chlorophenoxy acid herbicides in aqueous samples

A novel sample preparation method “Dispersive liquid–liquid–liquid microextraction” (DLLLME) was developed in this study. DLLLME was combined with liquid chromatography system to determine chlorophenoxy acid herbicide in aqueous samples. DLLLME is a rapid and environmentally friendly sample pretreatment method. In this study, 25 μL of 1,1,2,2-tetrachloroethane was added to the sample solution and the targeted analytes were extracted from the donor phase by manually shaking for 90 s. The organic phase was separated from the donor phase by centrifugation and was transferred into an insert. Acceptor phase was added to this insert. The analytes were then back-extracted into the acceptor phase by mixing the organic and acceptor phases by pumping those two solutions with a syringe plunger. After centrifugation, the organic phase was settled and removed with a microsyringe. The acceptor phase was injected into the UPLC system by auto sampler. Fine droplets were formed by shaking and pumping with the syringe plunger in DLLLME. The large interfacial area provided good extraction efficiency and shortened the extraction time needed. Conventional LLLME requires an extraction time of 40–60 min; an extraction time of approximately 2 min is sufficient with DLLLME. The DLLLME technique shows good linearity (r² ≥ 0.999), good repeatability (RSD: 4.0–12.2% for tap water; 5.7–8.5% for river water) and high sensitivity (LODs: 0.10–0.60 μg/L for tap water; 0.11–0.95 μg/L for river water).     

Flotation-assisted dispersive liquid–liquid microextraction method for preconcentration and determination of trace amounts of cobalt: Orthogonal array design

A simple, rapid, and efficient flotation-assisted dispersive liquid–liquid microextraction method was developed for preconcentration of trace amount of cobalt(II) ions. In this technique, a mixture of toluene and methanol (20: 80, v/v) was injected through the septum in the bottom of a narrow-bore tube containing cobalt solution. Afterwards, the fine droplets of extraction solvent were formed and cobalt (as 1-nitroso-2- naphtol complex) was collected on the surface of solution by aeration. The effect of different variables on the extraction efficiency of cobalt such as pH of solution, ligand concentration and injection volume was investigated using orthogonal array design. At optimum conditions, the calibration curve was linear over the range of 10–1000 μg/L. The detection limit, relative standard deviation and enrichment factor were 3 μg/L, 3.9% (n = 10) and 120, respectively. The developed method was successfully applied to the determination of cobalt in water and drug samples.     

Application of vortex-assisted supramolecular solvent liquid–liquid microextraction for trace determination of nitroaniline isomers

A novel, fast and efficient method for the analysis of nitroaniline isomers as model compounds was developed using vortex-assisted supramolecular solvent liquid–liquid microextraction (VA-SMS-LLME). A vortex mixer was used as the mixer in supramolecular solvent liquid–liquid microextraction, and it decreased the extraction time greatly. Several important parameters influencing extraction efficiency, such as the type and volume of extraction solvent, pH of sample, salt effect and extraction time, were optimised in detail. Under the optimal conditions, the enrichment factor was 133 for p-nitroaniline, 98 for m-nitroaniline and 115 for o-nitroaniline, and the limits of detection by HPLC were 0.3, 1.0 and 0.5 μg L^?1, respectively. Linearity with determination coefficient from 0.9981 to 0.9993 was evaluated using water samples spiked with the nitroanilines at fourteen different concentration ranging from 4 to 1000 μg L^?1. The ranges of intra-day and inter-day precision (n = 5) at 10 μg L^?1 of nitroanilines were 1.67–7.05% and 9.4–11.6%, respectively. The VA-SMS-LLME method was successfully applied for preconcentration of nitroanilines in environmental water samples.     

In-syringe magnetic-stirring-assisted liquid–liquid microextraction for the spectrophotometric determination of Cr(VI) in waters

A fully automated method for the determination of chromate is described. It is based on the selective reaction of Cr(VI) with diphenylcarbazide in acidic media to form a colored complex of Cr(III) with the oxidation product diphenylcarbazone. The reaction was performed within the syringe of an automatic burette containing a magnetic stirrer for homogenization of the sample and the required reagents. In-syringe stirring was made possible using a specially designed driving device placed around the syringe barrel to achieve a rotating magnetic field in the syringe, forcing the stirrer to spin. In a second step, the reaction mixture in the syringe was neutralized to allow in-syringe magnetic-stirring-assisted dispersive liquid–liquid microextraction of the complex into 125 μL of n-hexanol. After phase separation by droplet flotation over 30 s, the organic phase was propelled into a coupled spectrophotometric detection cell. The entire multistep procedure including in-system standard preparation was done within 270 s. The method was used for the analysis of natural waters, achieving average analyte recovery of 103 %, a limit of detection of 0.26 μg L^-1, and a repeatability of less than 4 % relative standard deviation.     

Development of a dispersive liquid–liquid microextraction (DLLME) method coupled with GC/MS as a simple and valid method for simultaneous determination of phthalate metabolites in plasma

Phthalates are ubiquitous environmental contaminants, and frequent biological monitoring of their metabolites (as toxic species of phthalates in human body) is highly recommended. A novel dispersive liquid–liquid microextraction (DLLME) coupled with Gas Chromatography–Mass Spectrometry (GC-MS) has been developed for the determination of seven phthalate metabolites in human plasma for the first time. Plasma proteins were efficiently precipitated by adding of 0.2mg dry trichloroacetic acid to 10 mL plasma samples, incubation and centrifuging. For DLLME, a mixture of extraction solvent (chlorobenzene, 10 µL) and dispersive solvent (acetonitrile, 750 µL) were rapidly injected into 5.0 mL aqueous sample for the formation of cloudy solution, the analytes in the sample were extracted into the fine droplets of chlorobenzene. After extraction, phase separation was performed by centrifugation and the enriched analytes in the sedimented phase were subjected to GC-MS analysis. All important parameters affecting DLLME performance were investigated and optimised. Under the optimum extraction condition, the method yields a linear calibration curve for all target analytes in the concentration range from 5 to 5000 ng mL^?1. The limits of detection and relative standard deviations for all phthalate metabolites were between 1.21–2.09 ng mL^?1 and 4.8–6.8%, respectively. This is a very simple, rapid and reproducible method, which requires low volume of sample and toxic solvents.     

Dispersive liquid–liquid microextraction combined with high-performance liquid chromatography-UV detection as a very simple,rapid and sensitive method for the determination of bisphenol A in water samples

Dispersive liquid–liquid microextraction (DLLME) coupled with high-performance liquid chromatography (HPLC)-UV detection was applied for the extraction and determination of bisphenol A (BPA) in water samples. An appropriate mixture of acetone (disperser solvent) and chloroform (extraction solvent) was injected rapidly into a water sample containing BPA. After extraction, sedimented phase was analyzed by HPLC-UV. Under the optimum conditions (extractant solvent: 142 μL of chloroform, disperser solvent: 2.0 mL of acetone, and without salt addition), the calibration graph was linear in the range of 0.5–100 μg L⁻¹ with the detection limit of 0.07 μg L⁻¹ for BPA. The relative standard deviation (RSD, n = 5) for the extraction and determination of 100 μg L⁻¹ of BPA in the aqueous samples was 6.0%. The results showed that DLLME is a very simple, rapid, sensitive and efficient analytical method for the determination of trace amount of BPA in water samples and suitable results were obtained.