J. Guiteras

Determination of pKa values of some hydroxylated benzoic acids in methanol-water binary mixtures by LC methodology and potentiometry

An accurate estimation of pK(a) values in methanol-water binary mixtures is very important for several separation techniques such as liquid chromatography and capillary electrophoresis that use these solvent mixtures. In this study, the pK(a) values of 11 polyphenolic acids have been determined in methanol-water binary mixtures (10%, 20% and 30% (v/v)) by potentiometry, liquid chromatography (LC) and LC-DAD methodology. The results show a similar trend for the pK(a) values of all the studied compounds, as they increase with increasing concentration of organic modifier, which allows a linear relationship between pK(a) values and mole fraction of methanol to be obtained. The pK(a) values obtained in aqueous medium have been compared with those given in the literature, and also with the values predicted by the SPARC on-line pK(a) calculator. The data obtained have been used to test the feasibility of an estimation of dissociation constants in a methanol-water medium from the relationship between pK(a) values and the organic cosolvent fraction in the mixtures.

Use of cloud point extraction methodology for the determination of PAHs priority pollutants in water samples by high-performance liquid chromatography with fluorescence detection and wavelength programming

The cloud point extraction methodology has been used to develop a new procedure for the preconcentration of polycyclic aromatic hydrocarbons in water samples (bottled and network supply). Triton X-114 was used as extractant agent. The procedure, consisting of three steps (preconcentration into the surfactant, clean-up and concentration of the eluate), allows good detection limits to be reached (from nanograms per liter to even sub-nanograms per liter in some cases). Determination was carried out by HPLC, with fluorimetric detection and wavelength programming.

Development and validation of a multiclass method for the analysis of antibiotic residues in eggs by liquid chromatography–tandem mass spectrometry

A multiclass method for the analysis of residues, in egg matrices, of 41 antimicrobial agents belonging to seven families (sulfonamides, diaminopyridine derivates, quinolones, tetracyclines, macrolides, penicillins and lincosamides) was developed and validated according to the requirements of European Commission Decision 2002/657. Compounds were extracted with a pressurized liquid extraction (PLE) technique using a 1:1 mixture of acetonitrile and a succinic acid buffer (pH 6.0) at 70 °C. As this resulted in clear extracts, no further clean-up was necessary. Analytes were determined by ultra-high-pressure liquid chromatography-tandem mass spectrometry (UHPL-MS/MS) in a chromatographic run of 13 min. Calibration was carried out with spiked blank samples subjected to the entire analytical procedure. Five compounds, two of them isotopically labelled, were used as internal standards. Most analytes were quantified with errors below 10%. Precision in terms of reproducibility standard deviation was between 10% and 20% in most cases. CCα values were in the range 0.5-3.8 μg kg⁻¹ for the non-authorized compounds. The proposed method would enable an experienced analyst to process about 25 samples per day.

Antibiotics in food: Legislation and validation of analytical methodologies

In this paper, European food safety legislation is presented, and special attention is devoted to monitoring residues of veterinary drugs in foodstuffs of animal origin. After a short review of the state of the art of analytical methodology for antibiotic residue analysis, the paper focuses on validation of analytical methods, with Decision 657/2002/EC as reference document. Finally, the main issues of the quality control of the analytical data, i.e. analysis of reference materials and participation in proficiency tests, are briefly addressed.

Spectrofluorimetric detection of zinc and cadmium with 8-(benzenesulfonamido)-quinoline immobilized on a polymeric matrix

A chelating resin, 8-(benzenesulfonamido)quinoline immobilized on an XAD-7 support, was prepared and its use for the spectrofluorimetric determination of Zn and Cd was investigated. The immobilized material was placed in a flow cell in a spectrofluorimeter and used in a flow injection system; the fluorescent complex was formed and immobilized in the cell. The chelating resin could be stored for at least three months and used repeatedly without any appreciable amount of reagent being lost. Calibration graphs were linear for up to 500 µg l–1 of Zn or Cd. Detection limits were 1.6 µg l–1 for Zn and 1.9 µg l–1 for Cd. Quantification limits were 5.5 µg l–1 for Zn and 6.8 µg l–1 for Cd. Precisions at 50 µg l–1 were 1.7% for Zn and 1.2% for Cd. Detection limits can be improved by injecting larger volumes of sample. Preconcentration factors of up to 10 were obtained for both metal ions.

Multivariate calibration applied to synchronous fluorescence spectrometry. Simultaneous determination of polycyclic aromatic hydrocarbons in water samples.

Synchronous fluorescence spectra of mixtures containing ten polycyclic aromatic hydrocarbons (anthracene, benz[a]anthracene, benzo[a]pyrene, chrysene, fluoranthene, fluorene, naphthalene, perylene, phenanthrene and pyrene) have been used for the determination of these compounds by Partial Least Squares Regression (PLSR), using both PLS-1 and PLS-2. Different procedures have been used for the pretreatment of the data in order to obtain better models, and the size of the calibration matrix has also been studied. The best models have been used for the determination of the above mentioned PAHs in spiked natural water samples at concentration levels between 4 and 20 ng ml(-1). Recoveries ranged from 80 to 120% in most cases, although fluorene gave significantly lower results.

Validation of a method for the analysis of nine quinolones in eggs by pressurized liquid extraction and liquid chromatography with fluorescence detection.

A multiresidue method for the analysis, in egg matrices, of residues of nine quinolones used in veterinary medicine, has been developed and validated according to the provisions of Council Decision 2002/657. Compounds were extracted by a pressurized liquid extraction (PLE) technique using a 1:1 mixture of acetonitrile and a phosphoric acid buffer (pH 3.0) at 70°C. The obtained extract was clear enough, so that no further clean-up was necessary. Analytes were determined by liquid chromatography (LC) with fluorescence detection (FL). Two chromatographic columns were compared: a high-purity silica Inertsil C(8) column and a newly developed Kinetex C(18) core-shell technology column. Validation was carried out at four concentration levels, using both chromatographic columns. Precision in terms of reproducibility standard deviation was between 7% and 23%, and good recoveries were obtained. Decision limit (CCα) and detection capability (CCβ) values obtained with the Inertsil and Kinetex columns were in the 0.2-19.8 μg kg(-1) and 0.4-33.5 μg kg(-1) concentration ranges, respectively. The proposed method allows residues of quinolones banned for use with laying hens to be detected and quantified in eggs. About twenty-four samples per day can be processed.

Development of fast-scanning fluorescence spectra as a detection system for high-performance liquid chromatography : Determination of polycyclic aromatic hydrocarbons in water samples

Abstract A new detection system based on full-spectrum fast scanning fluorescence has been developed for high-performance liquid chromatography (HPLC-FSFS). Emission spectra were obtained by scanning the effluent from a chromatographic column at 50 nm s −1 . The raw chromatograms were corrected by an interpolation method, because the fluorescence intensity measured for each consecutive wavelength corresponded to a different portion of the effluent. Quantification of samples was done by partial least squares (PLS). This procedure has been used for the determination of polycyclic aromatic hydrocarbons in water samples.

Multiclass method for antimicrobial analysis in animal feeds by liquid chromatography–tandem mass spectrometry

AbstractA rapid multiclass method that covers 50 antimicrobials from 13 different families in animal feeds was developed. Samples were extracted using a mixture of methanol, acetonitrile and a McIlvaine buffer combined with sonication. Feed extracts were simply diluted prior to injection, since the clean-up strategies that were tested, based on either solid-phase extraction or dispersive solid-phase extraction, were ineffective at minimizing matrix-related signal suppression/enhancement. Analysis was carried out by liquid chromatography coupled to tandem mass spectrometry using an electrospray ionization source operating in positive and negative modes. For the quantification, matrix-fortified standard calibration curves were used to compensate for matrix effects and losses in sample preparation. The method was validated in-house in pig, poultry and cattle feed matrices and showed satisfactory performance characteristics. Thus, the proposed approach was suitable for application in a routine high-throughput laboratory for the official control of feeds. FigureMulticlass method for antimicrobial analysis in animal feeds.

Determination of Polycyclic Aromatic Hydrocarbons by HPLC with Spectrofluorimetric Detection and Wavelength Programming

Abstract The sixteen polycyclic aromatic hydrocarbons (PAH) classified as priority pollutants by the EPA have been determined by HPLC, in isocratic conditions, with spectrofluorimetric detection and wavelength programming. The possibility of selecting the optimum wavelengths for each PAH gives the advantages of increased sensitivity - reaching the low pg level - and of improved selectivity, because it is possible to carry out an independent determination of coeluting compounds, provided that their excitation and emission spectra are different enough.