Marina F. M. Tavares

Investigation of preconcentration strategies for the trace analysis of multi-residue pesticides in real samples by capillary electrophoresis

In this work, on-line preconcentration strategies were investigated for the multi-residue analysis of pesticides in drinking water and vegetables using micellar electrokinetic chromatography. Among the on-line strategies, sweeping and stacking with reverse migration of micelles (SRMM), with and without the insertion of a plug of water before sample injection, were contrasted. A new version of SRMM was also introduced. The modification consisted of momentarily applying a positive voltage at the inlet vial right after sample has been injected, increasing the efficiency by which the analytes are captured. Nine pesticides from different classes, carbendazim (benzimidazole), simazine, atrazine, propazine and ametryn (triazine), diuron and linuron (urea), carbaryl and propoxur (carbamate), were baseline separated in less than 6 min with a electrolyte composed of 20 mmol l(-1) phosphate buffer at pH 2.5, containing 25 mmol l(-1) sodium dodecyl sulfate and 10% methanol. Limits of detection (LODs) in the order of 2-46 microg l(-1) for the pesticides under investigation were obtained solely using the on-line strategies. Enrichment factors of 3-18-fold were obtained. These factors were computed as the improvement of the concentration LODs with respect to the reference condition (injection of 10 s at 2.5 kPa pressure). The proposed methodologies were applied to the analysis of pesticides in complex matrices such as carrot extracts where the detection of 2.5 microg l(-1) was illustrated. By combining off-line solid-phase extraction and the proposed on-line strategies, the detection of pesticides in drinking water at the 0.1 microg l(-1) level was conceived.

Mecanismos de Separação em Eletroforese Capilar

Since its inception in the 80s, capillary electrophoresis has matured into a well established technique for the separation and analysis of complex samples. One of its strongest aspects is the ability to handle materials from a diversity of chemical classes, ranging from few to millions of Daltons. This is only possible because several modes of electrophoresis can be performed in a single capillary format. In this work, relevant aspects of capillary zone electrophoresis in its three modes (free solution, micellar and gel), capillary isoelectric focusing and capillary isotachophoresis are discussed and many representative applications are presented.

Modified electroosmotic flow by cationic surfactant additives in capillary electrophoresis Evaluation of electrolyte systems for anion analysis

A detailed experimental study of the variables affecting flow reversal was conducted in electrolyte systems commonly used for the determination of small anions. The electroosmotic flow mobilities of chromate (5 mM, pH 8) and 3,5-dinitrobenzoate solutions (concentration range 1.25–10 mM and pH 4–9) were determined by monitoring the migration time of pure water with indirect UV detection at 254 nm. The addition of the cationic surfactants cetyltrimethylammonium bromide, tetradecyltrimethylammonium bromide and lauryltrimethylammonium bromide, computed as fractions of the standard critical micelle concentration (std CMC), was investigated. The results suggested that flow reversal follows a single transition adsorption mechanism, in which the capillary inner surface is occupied by a layer of hemi-micelles, i.e. two dimensional aggregates formed by lateral interaction of surfactant tails. Once flow reversal is achieved, with surfactant concentration approximately above 10% std CMC, the nature of the electrolyte anion as well as the pH of the solution does not exert any significant effect on the magnitude of the electroosmotic flow. However, at this condition, the electroosmotic flow mobility varies linearly with the logarithm of the anion concentration.

Determination of amino acids by capillary electrophoresis‐electrospray ionization‐mass spectrometry: An evaluation of different protein hydrolysis procedures

In this work, a CE equipment, online hyphenated to an IT MS analyzer by a linear sheath liquid interface promoting ESI, was used to develop a method for quantitative determination of amino acids. Under appropriate conditions (BGE composition, 0.8% HCOOH, 20% CH3OH; sheath liquid composition, 0.8% HCOOH, 60% methanol; VESI, +4.50 kV), analytical curves of all amino acids from 3 to 80 mg/L were recorded presenting acceptable linearity (r >0.99). LODs in the range of 16–172 μmol/L were obtained. BSA, a model protein, was submitted to different hydrolysis procedures (classical acid and basic, and catalyzed by the H+ form of a cation exchanger resin) and its amino acid profiles determined. In general, the resin‐mediated hydrolysis yields were overall similar or better than those obtained by classical acid or basic hydrolysis. The resulting experimental‐to‐theoretical BSA concentration ratios served as correction factors for the quantitation of amino acids in Brazil nut resin generated hydrolysates.

Laser-induced fluorescence and UV detection of derivatized aldehydes in air samples using capillary electrophoresis

In this work, two capillary zone electrophoresis methodologies using UV absorption detection (214 nm) and laser-induced fluorescence detection (He/Cd laser, 325 nm excitation, 520 nm emission) of selected aldehydes (formaldehyde, acetaldehyde, propionaldehyde and acrolein) derivatized with dansylhydrazine (DNSH, 5-dimethylaminonaphthalene-1-sulfohydrazide) were proposed and validated. The aldehydes react with DNSH to form negatively charged molecules in methanolic medium. In both methodologies, nine DNSH-derivatives, including isomers of acetaldehyde, propionaldehyde and acrolein and two impurities were baseline separated in 20 mmol l(-1) phosphate buffer at pH 7.02, in less than 9 min. The limits of detection for the UV and LIF methodologies ranged from 1.1-9.5 microg l(-1) and 0.29-5.3 microg l(-1), respectively. The applicability of both methodologies to contemplate real samples was confirmed in the analysis of aldehyde-DNSH derivatives in indoor and outdoor air samples.

Systematic approach to the separation of mono- and hydroxycarboxylic acids in environmental samples by ion chromatography and capillary electrophoresis

Abstract Strategies for method development and optimization of conditions in ion chromatography (IC) and capillary electrophoresis (CE) are proposed. The approaches are based on well established theoretical models for analyte elution and rely on the inspection of effective capacity factor versus specific gradient ramp slope curves for IC and effective mobility versus pH curves for CE. Commonly used IC eluent systems, such as sodium hydroxide and sodium borate, and the CE electrolyte 3,5-dinitrobenzoic acid were investigated. A standard aqueous solution containing formate, acetate, α-hydroxyacetate (glycolate), propionate, α-hydroxypropionate (lactate), butyrate and β-hydroxybutyrate anions was used to test the analytical conditions indicated by the models. Among the IC eluent systems, borate presented a better separation performance than hydroxide. However, a complete resolution of all components in the standard mixture was not accomplished experimentally at the chosen gradient ramp slope of 0.05 mM/ml. The analyte pairs acetate/lactate and glycolate/butyrate, whose effective capacity factors differ by approximately 0.2 units, co-eluted at 0.17 and 0.24 resolution, respectively, and the retention time of the last eluting analyte was relatively long (20 min). Nevertheless, the IC method provided the best overall limit of detection (LOD; 0.016–0.082 mg/l). Under the optimized CE conditions, all seven components in the standard mixture were resolved satisfactorily in less than 7 min. The analyte pair β-hydroxybutyrate/butyrate presented the worst resolution, 0.45, and a difference in effective mobility of 1.6%. The CE methodology provided the best column efficiency, roughly a ten-fold improvement in terms of number of plates per meter over the IC method, but the limit of detection was comparatively poorer (0.050–0.36 mg/l). Both proposed IC and CE methodologies can be applied to the analysis of mono- and hydroxycarboxylic acids in samples of environmental interest, providing complementary information. The choice of the most appropriate method is a compromise between chemical composition of the sample and concentration level of the analytes under investigation. For instance, the classical co-elution of acetate/lactate that occurs in IC columns, can be solved by CE. But the CE method may lack sensitivity for these analytes, which compromises the analysis of certain real samples. The LOD for acetate and lactate as determined by IC is 0.036 and 0.082 mg/l, respectively, while for CE they increase to 0.11 and 0.20 mg/l. Under optimum conditions, the separation and identification of mono- and hydroxycarboxylic acids in an atmospheric particulate matter sample is illustrated by both techniques.

CE‐ESI‐MS metabolic fingerprinting of Leishmania resistance to antimony treatment

Metabolomics has become an invaluable tool to unveil biology of pathogens, with immediate application to chemotherapy. It is currently accepted that there is not one single technique capable of obtaining the whole metabolic fingerprint of a biological system either due to their different physical‐chemical properties or concentrations. In this work, we have explored the capability of capillary electrophoresis mass spectrometry with a sheathless interface with electrospray ionization (CE‐ESI‐TOF‐MS) to separate metabolites in order to be used as a complementary technique to LC. As proof of concept, we have compared the metabolome of Leishmania infantum promastigotes BCN 150 (Sb (III) IC50 = 20.9 μM) and its variation when treated with 120 μM of Sb(III) potassium tartrate for 12 h, as well as with its Sb(III) resistant counterpart obtained by growth of the parasites under increasing Sb(III) in a step‐wise manner up to 180 μM. The number of metabolites compared were of 264 for BCN150 Sb(III) treated versus nontreated and of 195 for Sb(III) resistant versus susceptible parasites. After successive data filtering, differences in seven metabolites identified in databases for Leishmania pathways, showed the highest significant differences, corresponding mainly to amino acids or their metabolite surrogates. Most of them were assigned to sulfur containing amino acids and polyamine biosynthetic pathways, of special relevance considering the deterioration of the thiol‐dependent redox metabolism in Leishmania by Sb(III). Given the low concentrations typical for most of these metabolites, the assay can be considered a success that should be explored for new biological questions.

Multi-analytical platform metabolomic approach to study miltefosine mechanism of action and resistance in Leishmania

AbstractMiltefosine (MT) (hexadecylphosphocholine) was implemented to cope with resistance against antimonials, the classical treatment in Leishmaniasis. Given the scarcity of anti- Leishmania (L) drugs and the increasing appearance of resistance, there is an obvious need for understanding the mechanism of action and development of such resistance. Metabolomics is an increasingly popular tool in the life sciences due to it being a relatively fast and accurate technique that can be applied either with a particular focus or in a global manner to reveal new knowledge about biological systems. Three analytical platforms, gas chromatography (GC), liquid chromatography (LC) and capillary electrophoresis (CE) have been coupled to mass spectrometry (MS) to obtain a broad picture of metabolic changes in the parasite. Impairment of the polyamine metabolism from arginine (Arg) to trypanothione in susceptible parasites treated with MT was in some way expected, considering the reactive oxygen species (ROS) production described for MT. Importantly, in resistant parasites an increase in the levels of amino acids was the most outstanding feature, probably related to the adaptation of the resistant strain for its survival inside the parasitophorous vacuole. Online Abstract Figureᅟ

Determination of sorbate and benzoate in beverage samples by capillary electrophoresis-Optimization of the method with inspection of ionic mobilities.

The aim of this study was to develop a fast capillary electrophoresis method for the determination of benzoate and sorbate ions in commercial beverages. In the method development the pH and constituents of the background electrolyte were selected using the effective mobility versus pH curves. As the high resolution obtained experimentally for sorbate and benzoate in the studies presented in the literature is not in agreement with that expected from the ionic mobility values published, a procedure to determine these values was carried out. The salicylate ion was used as the internal standard. The background electrolyte was composed of 25 mmol L(-1) tris(hydroxymethyl)aminomethane and 12.5 mmol L(-1) 2-hydroxyisobutyric acid, at pH 8.1. Separation was conducted in a fused-silica capillary (32 cm total length and 8.5 cm effective length, 50 microm I.D.), with short-end injection configuration and direct UV detection at 200 nm for benzoate and salicylate and 254 nm for sorbate ions. The run time was only 28s. A few figures of merit of the proposed method include: good linearity (R(2)>0.999), limit of detection of 0.9 and 0.3 mg L(-1) for benzoate and sorbate, respectively, inter-day precision better than 2.7% (n=9) and recovery in the range 97.9-105%. Beverage samples were prepared by simple dilution with deionized water (1:11, v/v). Concentrations in the range of 197-401 mg L(-1) for benzoate and 28-144 mg L(-1) for sorbate were found in soft drinks and tea.

Multivariant optimization, validation, and application of capillary electrophoresis for simultaneous determination of polyphenols and phenolic acids in Brazilian wines

A method for the simultaneous determination of the stilbene resveratrol, four phenolic acids (syringic, coumaric, caffeic, and gallic acids), and five flavonoids (catechin, rutin, kaempferol, myricetin, and quercetin) in wine by CE was developed and validated. The CE electrolyte composition and instrumental conditions were optimized using 2(7-3) factorial design and response surface analysis, showing sodium tetraborate, MeOH, and their interaction as the most influential variables. The optimal electrophoretic conditions, minimizing the chromatographic resolution statistic values, consisted of 17 mmol/L sodium tetraborate with 20% methanol as electrolyte, constant voltage of 25 kV, hydrodynamic injection at 50 mbar for 3 s, and temperature of 25 degrees C. The R(2) values for linearity varied from 0.994 to 0.999; LOD and LOQ were 0.1 to 0.3 mg/L and 0.4 to 0.8 mg/L, respectively. The RSDs for migration time and peak area obtained from ten consecutive injections were less than 2% and recoveries varied from 97 to 102%. The method was applied to 23 samples of inexpensive Brazilian wines, showing wide compositional variation.