Browsing by Author "Simó, Carolina"
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- A sensitive micellar electrokinetic chromatography-laser induced fluorescence method to analyze chiral amino acids in orange juices.
2002-09-19T15:39:41Z In this work a new method to detect the existence of chiral amino acids in orange juice is presented. The method employs â-cyclodextrins and micellar electrokinetic chromatography with laser-induced fluorescence (MEKC-LIF) to separate and detect L- and D-amino acids (L-aa and D-aa) previously derivatized with fluorescein isothiocianate (FITC). A systematic optimization of the chiral-MEKC conditions is done bringing about in less than 20 min a good separation of the main amino acids found in orange juice (i.e., Pro, Asp, Ser, Asn, Glu, Ala, Arg, and the nonchiral GABA, i.e., ç-aminobutyric acid). Using this procedure, the analysis time reproducibility for the 15 standard compounds (L-aa, D-aa, and GABA) has been determined to be better than 0.2% (n ) 5) for the same day and better than 0.7% (n ) 15) for three different days. Corrected peak area reproducibility is somewhat lower, providing values better than 3.3% (n ) 5) for the same day and 6.9% (n ) 15) for three different days. The limit of detection using this procedure was determined to be 0.86 attomoles for L-Arg. The optimized FITC derivatization method allows the easy and straightforward detection of amino acids in orange concentrates and juices (i.e., only centrifugation of diluted samples for 5 min is needed prior to their derivatization). D-Ala, D-Asp, D-Arg, and D-Glu were determined in orange juices and orange concentrates from different geographical origins using this new method. Moreover, the effect of different temperature treatments (50, 92, and 150 °C) on the content of D-aa in orange juice was evaluated.
- Análisis de polímeros sintéticos por electroforesis capilar.
2001-09-19T15:39:18Z En este artículo se presenta una revisión de los diferentes trabajos que han aparecido en la bibliografía sobre el análisis de polímeros empleando electroforesis capilar (CE). Para ello, se describen primero los principios de funcionamiento de esta nueva técnica analítica, la instrumentación, los diferentes modos en que puede ser utilizada y su comparación con otras técnicas analíticas de separación. A continuación se describen las aplicaciones más representativas de CE en el análisis de polímeros sintéticos.
- Analysis of antioxidants from orange juice using countercurrent supercritical fluid extration, micellar electrokinetic chromatography and reverse phase-HPLC .
2002-09-19T15:39:44Z Antioxidants from orange juice were determined by the combined use of countercurrent supercritical fluid extraction (CC-SFE) prior to reverse-phase liquic chromatography (RP-LC) or micellar electrokinetic chromatography (MEKC). The separation of antioxidants found in the SFE fractions was achieved by using a new MEKC method and a published LC procedure, both using diode array detection. The characterization of the different antioxidants was further done by LC-mass spectrometry. Advantages and drawbacks of LC and MEKC for analyzing the antioxidants found in the different orange extracts are discussed. Although LC yields higher peak area and slightly better reproducibility than MEKC, the latter technique provides information about the CC-SFE extracts in analysis times 7 times faster than by LC. This analysis advantage can be used for the quick adjustment of CC-SFE conditions, thus providing a fast way to obtain orange fractions of specific composition.
- Analysis of synthetic macromolecules by capillary electrophoresis.
2002-09-19T15:40:19Z The complexity of the chemical composition of synthetic macromolecules has increased the need for more reliable analytical methodologies for characterizing these materials. CE has emerged as a powerful analytical tool able to provide useful information about the chemical properties of these complex molecules. Interestingly, such information can be in some cases complementary to that provided by other classical techniques. Thus, in this revision it has been demonstrated that CE is being used in their different modes (FSCE, CGE, MEKC, etc.) to successfully face the tremendous diversity that can be found analyzing synthetic polymers. One of the main characteristics of CE is that this technique makes possible to develop uniquely tailored separation procedures to analyze synthetic macromolecules of very different nature.
- Application of stepwise discriminat analysis to classify commercial orange juices... : some applications in food analysis.
2004-09-19T15:40:13Z The use of chiral amino acids content and stepwise discriminant analysis to classify three types of commercial orange juices (i.e., nectars, orange juices reconstituted from concentrates, and pasteurized orange juices not from concentrates) is presented. Micellar electrokinetic chromatography with laser-induced fluorescence (MEKC-LIF) and b-cyclodextrins are used to determine L- and D-amino acids previously derivatized with fluorescein isothiocyanate (FITC). This chiral MEKC-LIF procedure is easy to implement and provides information about the main amino acids content in orange juices (i.e., L-proline; L-aspartic acid, D-Asp, L-serine, L-asparagine, L-glutamic acid, D-Glu, L-alanine, L-.arginine, D-Arg, and the non-chiral g-amino-n-butyric acid (GABA), i.e., g-aminobutyric acid). From these results, it is clearly demonstrated that some D-amino acids occur naturally in orange juices. Application of stepwise discriminant analysis to 26 standard samples showed that the amino acids L-Arg, L-Asp and GABA were the most important variables to differentiate the three groups of samples. With these three selected amino acids a 100% correct classification of the samples was obtained either by standard or by leave-one-out cross-validation procedures. These classification functions based on the content in L-Arg, L-Asp and GABA were also applied to nine test samples and provided an adequate classification and/or interesting information on these samples. It is concluded that chiral MEKC-LIF analysis of amino acids and stepwise discriminant analysis can be used as a consistent procedure to classify commercial orange juices providing useful information about their quality and processing. To our knowledge, this is the first report about the combined use of chiral capillary electrophoresis and discriminant techniques to classify foods.
- Chiral capillary electrophoresis-mass spectrometry of amino acids in foods.
2005-09-19T15:39:51Z In this work, the development of a new chiral capillary electrophoresis-mass spectrometry (CE-MS) method to separate D- and L-amino acids is shown. On-line coupling between CE and MS is established through an electrospray-coaxial sheath flow interface. Enantiomer separation is achieved by using a cheap, nonvolatile, chiral selector as b-cyclodextrin in the background electrolyte (BGE) together with a physically coated capillary that is aimed to prevent contamination of the electrospray. The capillary coating is simple and easy to obtain as it only requires flushing of the capillary with a polymer aqueous solution for 3 min. Optimization of CE parameters (pH of BGE, type and concentration of chiral selector, and capillary inner diameter) and electrospray-MS parameters (nature and flow rate of the sheath liquid, nebulizer pressure) is carried out. Two different derivatization protocols of amino acids using dansyl chloride (DNS) and fluorescein isothiocyanate (FITC) are compared in terms of MS sensitivity and chiral resolution. Under optimum CE-MS conditions it is observed that the MS sensitivity obtained for FITC- and DNS-amino acids is similar (with limit of detection (LOD) in the mM range, corresponding to amounts injected in the fmol range) while chiral resolution is better for FITC-amino acids. The optimized method is demonstrated to provide the simultaneous analysis of 15 selected amino acids (i.e., FITC-D/L-Asp, -Glu, -Ser, -Asn, -Ala, -Pro, -Arg, and FITC-g-aminobutyric acid (GABA) in a single chiral CE-MS run, corresponding to the main amino acids that can be found in orange. Moreover, as a result of the high resolution achieved, it is possible to detect down to 2% of D-Asp in the presence of 98% of L-Asp. The good possibilities of chiral CE-MS in food analysis are corroborated through the detection of the main amino acids in a commercial orange juice (i.e., FITC-L-Asp, -Glu, -Ser, -Asn, -Pro, -Arg, and the nonchiral FITCGABA) as well as the determination of the fraudulent addition of synthetic amino acids (containing D- and L-forms) to a fresh orange juice
- Chiral capillary electrophoresis-mass spectrometry of food analysis.
2005-09-19T15:39:52Z A new physically adsorbed capillary coating for capillary electrophoresis-mass spectrometry (CE-MS) of basic proteins is presented, which is easily obtained by flushing the capillary with a polymer aqueous solution for two min. This coating significantly reduces the electrostatic adsorption of a group of basic proteins (i.e., cytochrome c, lysozyme, and ribonuclease A) onto the capillary wall allowing their analysis by CE-MS. The coating protocol is compatible with electrospray inonlzation (ESl)-MS via the reproducible separation of the standard basic proteins (%RSD values (n = 5) < 1 % for analysis time reproducibility and < 5% for peak heights, measured from the total ion electropherograms (flEs) within the same day). The LODs determined using cytochrome c with total ion current and extracted ion current defection were 24.5 and 2.9 fmol, respectively. Using this new coating lysozymes from chicken and turkey egg white could be easily distinguished by CE-MS, demonstrating the usefulness of this method to differentiate animal species. Even after sterilization at 1200C for 30 min, lysozyme could be detected, as well as in wines at concentrations much lower than the limit marked by the EC Commission Regulation. Adulteration of minced meat with 5% of egg-white could also be analysed by our CE-MS protocol.
- Chiral electromigration methods in food analysis.
2003-09-19T15:40:12Z This review article addresses the different chiral capillary electrophoretic methods that are being used for the study and characterization of foods and food compounds (e.g., amino acids, organic acids, sugars, pesticides). An updated overview, including works published till December 2002, on the principal applications of enantioselective procedures together with their main advantages and drawbacks in food analysis is provided. Some anticipated applications of chiral electromigration methods in food characterization are also discussed.
- Combining peptide modeling and capillary electrophoresis-mass Spectrometry for Characterization of Enzymes Cleavage Patterns: Recombinant versus Natural Bovine Pepsin A.
2005-09-19T15:40:23Z Nowadays there is an increasing number of recombinant enzymes made available to industry. Before replacing the use of natural enzymes with their cognate recombinant counterparts, one important issue to address is their actual equivalence. For a given recombinant proteolytic enzyme, its equivalence can be investigated by comparing its cleavage specificity with that obtained from the natural enzyme. This is mostly done by analyzing the fragments (i.e., peptidic map) attained after enzymatic digestion of a given protein used as substrate. The peptidic maps obtained are typically characterized using separation techniques together with MS and MS/MS systems. However, these procedures are known to be difficult and laborintensive. In this work, the combined use of a theoretical model that relates electrophoretic behavior of peptides to their sequence together with capillary electrophoresismass spectrometry (CE-MS) is proposed to characterize in a very fast and simple way the cleavage specificity of new recombinant enzymes. Namely, the effectiveness of this procedure is demonstrated by analyzing in few minutes the fragments obtained from a protein hydrolysated using recombinant and natural pepsin A. The usefulness of this strategy is further corroborated by CEMS/ MS. The proposed procedure is applicable in many other proteomic studies involving CE-MS of peptides.
- Detection and quantitation of a bioactive compound in vicia narbonensis L. seeds by capillary electrophoresis-mass spectrometry: a comparative study with UV detection.
2005-09-19T15:39:21Z Capillary zone electrophoresis with mass spectrometry (CE-MS) and UV detection (CE-UV) was applied to the quantitative determination of g-glutamyl-S-ethenyl-cysteine (GEC), a bioactive and unstable compound present in Vicia narbonensis L. seeds. This compound is responsible for, among other negative effects, palatability reduction and grain toxicity. In order to carry out the quantitative analysis of GEC, different conditions (such as composition, concentration and pH of the background electrolyte, and type and time of extraction) were studied. Also, adequate conditions for electrospraymass spectrometry of this bioactive compound were investigated. The best extraction conditions of GEC from V. narbonensis L. seeds flour were obtained using ethanolwater (70:30 v/v) for 45 min. The use of a 20 m ammonium hydrogen carbonate at pH 7 provided adequate analytical conditions compatible with the unstable nature of GEC as well as with the requirements of CE-UV and CE-MS analysis. A comparative study was carried out between the different figures of merit of CE-UV and CE-MS for quantitative purposes. Both techniques provided similar limit of detection and can be applied with confidence within the same linear dynamic range. However, reproducibility and speed of analysis were better using CE-UV. The developed methods were readily applied to quantify GEC in seeds of 21 genotypes of V. narbonensis L.. A good agreement between CE-MS and CE-UV results was observed corroborating the usefulness of both approaches for quantitative purposes.
- Evaluation of filter paper collection of urine samples for detection and measurement of organic acidurias by capillary electrophoresis.
2002-09-19T15:39:35Z There is little doubt that mental retardation has been prevented in most babies diagnosed by newborn screening programs for inborn errors and the cost–benefit ratios of these programs have been reported as highly positive. In a previous work we optimised a CE method for quick profiling of organic acidurias, which characterize a large number of inborn errors, so that it permits the separation, detection and even identification in less than 15 min of 22 organic acids in urine samples related to a wide range of metabolic disorders. In the present work we have studied the adequacy of filter paper collection of urine samples to simplify this step, always difficult in babies, when it is not performed by training personnel. The studied parameters were: media and conditions for re-extraction to give the best sensitivity and a more simple procedure when the samples are measured by CE, interferences coming from the diaper, recoveries obtained, possible correction of recoveries with creatinine and stability of the compounds. The whole method we report has the advantages of easy sample collection, easy shipping or delivery, and rapid analysis. Moreover, this method of collection and analysis allows the identification and quantitation of fumaric, methylmalonic, N-acetylaspartic, pyroglutamic and homogentisic acids, as well as glutaric acid for which screening is considered especially advisable.