A new method based on dispersive solid phase microextraction with commercial MOFs coupled to LC-MS/MS for the determination of isoflavones in soy drinks.

For the first time, a method based on dispersive solid phase microextraction (D-µSPE) using commercial metal-organic frameworks coupled to liquid chromatography-triple quadrupole tandem mass spectrometry (LC-MS/MS) has been proposed for the determination of isoflavones in soy drinks. The use of commercial sorbents simplifies the sample treatment procedure and allows their application to routine analysis. Optimization of the parameters involved in the microextraction process was carried out using a Box-Behnken experimental design. Under the optimized conditions, the limits of detection ranged between 2 and 7 µg L-1; the intra-day and inter-day precision were <10 and 20%, respectively, and the recoveries were in the range of 61-120%. No significant matrix effect was found, which allowed the use of external standard calibration method. The method was successfully applied to the determination of isoflavones in commercial soy milk samples.

A confirmatory method for the determination of phenolic endocrine disruptors in honey using restricted-access material-liquid chromatography-tandem mass spectrometry.

The present work describes the development and validation of an analytical method based on liquid chromatography (LC), coupled with tandem mass spectrometry (MS/MS) that allows the determination and confirmation of several endocrine-disrupting chemicals (EDCs) in honey. The EDCs studied were nine phenols of different nature: chlorophenols (2,4-dichlorophenol, 2,4,5-trichlorophenol, and pentachlorophenol), alkylphenols (4-tert-butylphenol, 4-tert-octylphenol, and 4-n-octylphenol) bisphenols (bisphenol-A and bisphenol-F), and 4-tert-butylbenzoic acid. The method incorporates a restricted-access material (RAM), coupled on-line to the LC-MS/MS system, which allows direct injection of the matrix into the RAM-LC-MS/MS system. The optimized method developed, RAM-LC-MS/MS, was applied to fortified honey samples, affording detection limits in the 0.6-7.2 ng g(-1) range, calculated for a signal-to-noise ratio of 3. In addition, the method was validated as a quantitative confirmatory method according to European Union Decision 2002/657/EC. The validation criteria evaluated were linearity, repeatability, reproducibility, recovery, decision limits, detection capabilities, specificity, and ruggedness. Repeatability and within-laboratory reproducibility were evaluated at two concentration levels, being +/-11% or below at 20 ng g(-1). The decision limits (CC(alpha)) and detection capabilities (CC(beta)) were in the 1.7-12.6 and 2.8-21.6 ng g(-1) range, respectively.

Determination of endocrine-disrupting compounds in cereals by pressurized liquid extraction and liquid chromatography-mass spectrometry. Study of background contamination.

A sensitive method based on pressurized liquid extraction (PLE) and liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) has been developed for the determination in cereal samples of seven endocrine-disrupting compounds: bisphenol A (BPA), 4-tert-butylbenzoic acid (BBA), 4-nonylphenol (NP), 4-tert-butylphenol (t-BP), 2,4-dichlorophenol (DCP), 2,4,5-trichlorophenol (TCP) and pentachlorophenol (PCP). For the PLE procedure, methanol was selected as the extraction solvent. An experimental design approach was applied to optimize other PLE parameters. The recoveries achieved for the all seven compounds were in the 81-104% range, with relative standard deviations of 4-9%. An additional preconcentration step, based on solid-phase extraction (SPE), after the PLE step proved to be a successful way for obtaining a more sensitive method. The detection limits achieved in corn breakfast cereals were in the 0.003-0.013 microg g(-1) range, except for BPA, with a detection limit of 0.043 microg g(-1), for a sample size of 2.5 g. These values are similar to or even lower than currently legislated limits for pesticides in cereals and cereal-based foodstuffs. We also investigated possible contamination during the experimental process by the target compounds released from purified water, plastics, syringes, peristaltic pump tubes, glassware and other laboratory materials in contact with the samples along the analytical process.

Comparison of a non-aqueous capillary electrophoresis method with high performance liquid chromatography for the determination of herbicides and metabolites in water samples.

A method of capillary electrophoresis (CE) for the determination of triazine herbicides and some of their main metabolites in water samples has been developed. The proposed CE method includes an off-line solid-phase extraction (SPE) procedure with LiChrolut EN sorbent coupled to a non-aqueous capillary electrophoresis (NACE) separation with UV detection. The target compounds were the chloro-s-triazines simazine, atrazine, propazine; the methyltio-s-triazines ametryn and prometryn and three main derivatives from the atrazine degradation products; namely, deethylatrazine, deethylhydroxyatrazine and deisopropylhydroxyatrazine. The analytical characteristics of the CE method are reported. The repeatability of the method was studied considering the different steps of the method separately in order to determine the contributions of each step to the total variability of the method. The NACE-UV results are compared with those obtained with a high performance liquid chromatography with UV detection (HPLC-UV) method. The same off-line SPE procedure was applied to both techniques. The results obtained show that both methods afford the same results in the analysis of surface and drinking water samples, with a level of significance regarding the F- and t-tests greater than 0.05 in all the cases. The detection limits in surface water samples were in the 0.04-0.32 microg l(-1) and 0.11-1.2 microg l(-1) ranges for the NACE-UV and HPLC-UV methods, respectively. The recoveries (spiked/found) were significantly 100% in all cases.

Determination of triazines and dealkylated and hydroxylated metabolites in river water using a propazine-imprinted polymer.

A molecularly imprinted polymer (MIP) obtained by precipitation polymerisation using propazine as template has been employed as sorbent for the solid phase extraction of triazines and some of their hydroxylated and dealkylated metabolites from river water. Three configurations were studied: (a) use of the propazine-MIP as a selective sorbent for the extraction of triazines directly from water; (b) use of mixtures of LiChrolut EN (a polymeric sorbent of styrene divinylbenzene) and propazine-MIP as sorbent, and (c) use of propazine-MIP as a clean-up sorbent for organic extracts obtained in a prior SPE procedure with LiChrolut EN. The former two configurations imply that the analytes pass through the propazine-MIP in aqueous medium, whereas in the latter case the analytes percolate through the propazine-MIP in an organic medium coming from the previous SPE step. Different types of water were tested to assess matrix effects. The analytical characteristics of the three configurations were evaluated.

Pressurized liquid extraction in the analysis of food and biological samples.

Originally, the use of the pressurized liquid extraction technique (PLE) was mainly focused on the extraction of environmental pollutants present in soil matrices, sediments, and sewage sludge. However, more recently the distinct advantages of this technique are being exploited in diverse areas, including biology, and the pharmaceutical and food industries. The aim of the present review is to explore recent analytical applications of this extraction technique (PLE) in the extraction of contaminant compounds and matrix components in food and biological samples, placing special emphasis on the strategies followed to obtain a rapid, selective, efficient and reliable extraction process.

Determination of weakly acidic endocrine-disrupting compounds by liquid chromatography-mass spectrometry with post-column base addition.

A sensitive analytical method based on liquid chromatography-electrospray ionisation mass spectrometry (LC-ESI-MS) has been developed for the determination of seven endocrine-disrupting compounds: 4-n-nonylphenol (NP), 4-tert-butylphenol (t-BP), bisphenol A (BPA), 2,4-dichlorophenol (DCP), 2,4,5-trichlorophenol (TCP), pentachlorophenol (PCP) and 4-tert-butylbenzoic acid (BBA) in water samples. To achieve a good LC separation, acidification of the LC mobile phase was necessary, but this led to MS signal suppression for the less acidic compounds. In order to enhance the sensitivity for these analytes, post-column addition of different bases such as ammonia, trimethylamine, and 1,8-diazabicyclo-(5,4,0)undec-7-en (DBU) was evaluated. The post-column addition of base is proposed here to raise effluent pH, helping in the ionisation process of the compounds with higher pKa values (t-BP, BPA, DCP and NP). The use of DBU, diluted in MeOH, proved to be the most efficient post-column reagent for enhancing the MS signal. The signal-to-noise ratios for t-BP and NP increased by more than 200-fold and 35-fold, respectively, whereas for DCP and BPA an increase of about 10-fold was achieved. This strategy permitted direct determination of the seven compounds at low ppb levels. For application to real water samples, an extraction and preconcentration step using the solid-phase extraction (SPE) technique was carried out. The applicability of three solid-phase materials--Bond Elut C18, and two polymeric sorbents: LiChrolut EN and Oasis HLB--and the optimization of other SPE parameters such as the elution solvent and sample volume used, were studied in order to maximize extraction efficiency. Oasis HLB provided the best results, obtaining--with the proposed SPE procedure--satisfactory percentage recoveries for all compounds (70-110%) with the exception of NP, for which a recovery of 54% was achieved. Application of the whole method, SPE-LC-(ESI)-MS, to natural waters permitted low nanogram-per-liter determination of all seven compounds.

Prediction of the behaviour of organic pollutants using cloud point extraction.

A preconcentration study based on the cloud point phenomenon was carried out for a set of triazine herbicides, three of them chloro-substituted and three of them methylthio-substituted. Concentration factors and recoveries were calculated as function of the percentage of the non-ionic surfactant Triton X-114 employed. From these values, obtained from a cloud point extraction (CPE) procedure, the distribution coefficient between the Triton X-114 micelles and water, Kc, prior to CPE was calculated for each triazine and related to the corresponding octanol-water partition coefficient, Kow. In order to confirm the results obtained with the triazine herbicides, two sets of data from chemically different organic pollutants--organophosporous and chlorophenols--obtained from the literature were assessed, concluding that they display a similar behaviour to that of the triazine herbicides. This can be used to predict the CPE behaviour of other organic pollutants from their octanol-water partition coefficients. The Kc values were compared with the analyte concentration ratio in the surfactant-rich phase and aqueous phase (Ksa) with a view to obtaining a link between the analyte behaviour prior to and after cloud point extraction procedures.

Determination of herbicides, including thermally labile phenylureas, by solid-phase microextraction and gas chromatography-mass spectrometry.

A method for the determination of 10 herbicides, including thermally unstable compounds, has been developed. The method uses solid-phase microextraction (SPME) with a polyacrylate fibre. Separation, identification and quantification were accomplished with gas chromatography-mass spectrometry. The herbicides chosen belong to different chemical groups and were alachlor, atrazine, chlorotoluron, diclofop, diflufenicam, ethofumesate, isoproturon, linuron, terbutryn and trifluralin. In the present work we studied the chromatographic behaviour of three phenylureas as a function of the medium and injection mode employed. The compounds generated as a function of the solvent used in direct injection of the phenylureas (ethyl acetate, methanol and methanol-water) and those obtained when injection was accomplished using the polyacrylate fibre were determined. The results allow us to propose a method for the determination of stable and thermally unstable herbicides as long as a preconcentration step involving SPME is carried out. In the proposed method, the limits of detection varied between 0.02 microg/l for ethofumesate and 0.11 microg/l for chlorotoluron. The method was applied to the determination of these herbicides in surface and ground water samples, performing quantification by standard addition calibration. The contents of chlorotoluron and atrazine found were significantly equal to those obtained using HPLC after a preconcentration stepwith styrene-divinylbenzene sorbents.

Analysis of pesticide residues in matrices with high lipid contents by membrane separation coupled on-line to a high-performance liquid chromatography system.

Separation through membranes coupled to an HPLC system was used as a technique for the analysis of pesticide multiresidues in samples with high lipid contents. As well as the usual procedure, in the proposed system it is possible to recirculate the sample through the membrane cell, which permits the extraction system to be applied to cases in which only a very small volume of sample is available. A procedure for pesticide multiresidue analysis in egg samples was developed as a representative example of the applicability of the proposed method. To accomplish this, the analytes (dichlorvos, dimethoate, propoxur, paraoxon, pirimicarb, atrazine, ametryne, terbutryne, azinphos-methyl, folpet) were subjected to prior extraction in a Soxhlet system, after which the extract was introduced into the membrane separation device coupled to the HPLC system. This procedure afforded clean chromatograms, hence considerably facilitating determination, and at the same time was efficient in removing macromolecular compounds. For egg samples, spiked at a concentration level of 0.750 mg/kg, recoveries ranged from 60 to 98%. The detection limits varied from 0.018 mg/kg for dichlorvos to 0.002 mg/kg for atrazine.

Determination of triazine herbicides in natural waters by solid-phase extraction and non-aqueous capillary zone electrophoresis.

Capillary zone electrophoresis (CZE) in an organic medium was used to analyse triazines at sub-ppb concentration levels in natural waters after a preconcentration step using conventional C18 cartridges and new Oasis HLB devices. With both sorbents, satisfactory results were obtained on analysing deionized water. However, on analysing natural waters, both sorbents showed very different types of behaviour. The different variables affecting the elution of both sorbents were studied, resulting in the choice of Oasis HLB as the most suitable for later separation by CZE in non-aqueous medium. Combination of a preconcentration step with electrokinetic injection revealed that capillary electrophoresis with simple UV detection can also be used satisfactorily for the quantification of micropollutants in natural waters. The detection limits obtained varied between 0.01 and 0.05 microg l(-1), depending on the type of matrix analysed. The day-to-day precision varied between 0.9% and 2.3%, expressed as the relative standard deviation.

Solid-phase extraction and sample stacking-micellar electrokinetic capillary chromatography for the determination of multiresidues of herbicides and metabolites.

Micellar electrokinetic capillary chromatography (MEKC) with diode array detection was used for the separation of 13 compounds (eight herbicides widely used in agriculture: metribuzin, lenacil, ethofumesate, atrazine, terbutryn, isoproturon, chlorotoluron and linuron, and five of their principal degradation products; namely, deethylatrazine, 2-hydroxyatrazine, deethyl-2-hydroxyatrazine, deisopropylatrazine and 3-chloro-4-methylphenylurea). Peak separation for the 13 analytes was not successful when a single surfactant system was employed, neither sodium dodecyl sulfate (SDS) nor dioctyl sulfosuccinate (DOSS) sodium salt. However, a mixture of these herbicides was successfully separated using a mixed micellar system involving SDS-DOSS in less than 14 min. An application study of an on-line concentration technique for MEKC was carried out to enhance sensitivity. The optimized on-line stacking procedure consisted simply of the addition of 50 mM of sodium chloride to the injection sample, the stacking effect being more intensive as analyte polarity increased. When this stacking procedure was combined with an off-line sample preconcentration step, based on solid-phase extraction, analytes could be detected in the ppb range. The whole method was applied to ultra-high-quality and natural waters. Linear relationships between the analytical signal and the initial analyte concentration were found to be independent of the type of water, except for the more polar analytes for which small differences were observed.

Surfactant cloud point extraction and preconcentration of organic compounds prior to chromatography and capillary electrophoresis.

The use of preconcentration steps based on phase separation by the cloud point technique offers a convenient alternative to more conventional extraction systems. It has been used successfully for the preconcentration of species of widely differing character and nature, such as metal ions, proteins and other biomaterials, or organic compounds of strongly differing polarity. Here we address the most recent analytical applications of this methodology when used as an isolation and trace enrichment step prior to the analysis of organic compounds (polycyclic aromatic hydrocarbons, polychlorinated compounds, pesticides, phenolic derivatives, aromatic amines, vitamins, etc.) via liquid and gas chromatography or capillary electrophoresis.

Evaluation of surface- and ground-water pollution due to herbicides in agricultural areas of Zamora and Salamanca (Spain).

The pollution of agricultural land due to herbicides was assessed in the Guareña and Almar river basins, situated in the provinces of Zamora and Salamanca (Spain). A set of fifteen herbicides, including triazines, ureas, amides and others, was selected owing to their frequency of use, the amounts used, their toxicity and their persistence in the environment. Solid-phase extraction with polymeric cartridges, followed by HPLC with diode-array detection, were used to monitor the herbicides. This technique was chosen owing to the wide range of functionality and polarity of the analytes under study. The detection limits obtained were in the 0.004-0.025 microg/l range (lambda=220 nm). Surface and ground waters, taken from different locations in the basins, were analyzed over a 6-month period. The presence of six out of the fifteen herbicides monitored--chlortoluron, atrazine, terbutryn, alachlor, diflufenican and fluazifop-butyl--was detected in several samples at levels ranging from the detection limit to 1.2 microg/l. The relationship of these herbicides to the agricultural activities of the zone is discussed.

Analytical applications of membrane extraction in chromatography and electrophoresis.

An overview of the analytical applications of membrane-based systems for sample enrichment in chromatography and capillary electrophoresis is presented. A brief introduction to the different types of membranes and the main forces related to the transport through them is also given.

Development and validation of a method for the detection and confirmation of biomarkers of exposure in human urine by means of restricted access material-liquid chromatography-tandem mass spectrometry.

The present article describes the development and validation of a LC-MS/MS method for the determination and confirmation of biomarkers of exposure to different types of xenobiotics in human urine. The method combines the use of a restricted access material (RAM) coupled on-line to a LC-IT-MS system; in this way, a rapid and efficient matrix cleanup was achieved, reducing manual sample preparation to freezing and sample filtration. The ion trap (IT) mass spectrometry detector provided the selectivity, sensitivity and ruggedness needed for confirmatory purposes. The on-line RAM-LC-MS/MS method developed here has been validated as a quantitative confirmatory method according to the European Union (EU) Decision 2002/657/EC. The validation steps included the verification of linearity, repeatability, specificity, trueness/recovery, reproducibility, stability and ruggedness in fortified urine samples. Repeatability and within-laboratory reproducibility, measured as intraday and interday precisions, were evaluated at two concentration levels, being 12.7% or below at the concentration corresponding to the quantification limits. Matrix effects and non-targeted qualitative analyses were also evaluated in fortified urine samples. Decision limits (CC(alpha)) and detection capabilities (CC(beta)) were in the range of 3.6-16.5 and 6.0-28.1ngmL(-1) respectively. The results of the validation process revealed that the proposed method is suitable for reliable quantification and confirmation of biomarkers of exposure to xenobiotics in human urine at low ngmL(-1) levels. In addition, working in Data-Dependent Scan mode the proposed method can be used for the screening of these compounds in urine samples.

Use of a bisphenol-A imprinted polymer as a selective sorbent for the determination of phenols and phenoxyacids in honey by liquid chromatography with diode array and tandem mass spectrometric detection.

An extraction-preconcentration procedure based on the use of a molecularly imprinted polymer (MIP) as selective sorbent has been developed for the determination of several phenolic compounds (bisphenol-A, bisphenol-F and 4-nitrophenol) and phenoxyacid herbicides (2,4-D, 2,4,5-T and 2,4,5-TP) in honey samples. Liquid chromatography with diode array detection (LC-DAD) and electrospray ionisation-ion trap mass spectrometry (LC-IT-MS) were used for the separation, identification and quantification of these analytes. The molecularly imprinted polymer was obtained by precipitation polymerisation with bisphenol-A (BPA) as template and 4-vinylpyridine as the functional monomer. The behaviour of this sorbent was compared with those of other materials frequently used in SPE. The selectivity of the BPA-MIP for the target analytes was tested in samples containing other pesticides in common use. The recoveries achieved for all six compounds were in the 81-96% range. By applying the proposed procedure prior to LC-IT-MS, the limits of detection achieved in commercial honey samples were in the 0.1-3.8 ng g(-1) range, with relative standard deviations of 12-24%.

Development of a chemometric correlation technique to estimate acid-base descriptors for cationic acids in non-aqueous media.

We propose a procedure for estimating acid-base constants in organic solvents or mixture of solvents from the corresponding pK(a) values in aqueous medium and from certain properties of the organic solvents that characterize them. To accomplish this, we developed and validated a chemometric correlation for the calculation of the acid-base constants of different cationic acids in a broad variety of non-aqueous solvents. The parameters chosen for building the model were as follows: the acid-base constant of the compound in aqueous medium and those corresponding to the polarity-polarizability, basicity and acidity scales of the solvent. The results of the fitting were significant (p<0.01), with a root mean error in cross-validation of 18%, with no overfitting. The prediction of the acid-base constants for an external set of compounds had a mean absolute prediction error value of less than 0.8 pK(a) units.

Capillary zone electrophoresis in nonaqueous solvents in the presence of ionic additives.

Capillary zone electrophoresis (CZE) in nonaqueous media and in the presence of ionic additives has been successfully applied to the determination of compounds that differ only slightly in their electrophoretic mobilities. Triazine herbicides of environmental interest were chosen as test compounds because they behave as very weak bases. CZE separation of these analytes (especially chlorotriazines) in aqueous solution is difficult due to the low pH required for their conversion into protonated cationic form (HA(+)). However, in mixed nonaqueous solvents, 50% (v/v) acetonitrile-methanol, the acid-base characteristics of these compounds are modified, yielding the protonated ionic species that is susceptible to migration when subjected to an electric field. A noteworthy increase in separation selectivity and resolution can be achieved by using ionic additives. Thus, in this mode of capillary zone electrophoresis, separation is based on ionic interactions between the charged analytes and the ionic additive present in the separation medium. These interactions contribute to enhancing mobility differences and to improving analyte separation. For the separation of chloro- and methylthiotriazines, 10 mM perchloric acid in 50% (v/v) acetonitrile-methanol and 20 mM SDS proved to be satisfactory, providing high resolution in short analysis times. The selectivity achieved was found to depend on the degree of association of the analyte with the ionic additive in the nonaqueous medium. This permits manipulation of the selectivity of the electrophoretic separations as a function of the type and concentration of the ionic additive and of the nature of the nonaqueous medium employed.

Cloud point extraction as a preconcentration step prior to capillary electrophoresis.

Cloud point extraction was applied as a preconcentration step prior to capillary electrophoresis. The behavior of a surfactant-rich micellar phase injected into a capillary electrophoresis system was studied using different separation modes: micellar electrokinetic capillary chromatography and capillary zone electrophoresis (CZE). A problem that appeared on introducing a surfactant-rich phase into a bare fused silica capillary was that the surfactant was adsorbed onto the wall of the capillary, leading to a marked loss of efficiency and reproducibility both in the migration times and in the areas of the electrophoretic peaks. The use of cetyltrimethylammonium bromide dynamically coated capillaries afforded reproducible results, although the half-life of the capillary was short. The most satisfactory results were obtained by using nonaqueous media in the CZE mode, thus avoiding surfactant adsorption. Other parameters related to the composition of the injection medium were also studied to optimize the electrophoretic behavior of the analytes and the sensitivity of the determination. The optimized procedure was applied to the determination of triazines in tap and river water samples.