Triacylglycerols Determination by High-temperature Gas Chromatography in the Analysis of Vegetable Oils and Foods: A Review of the Past 10 Years.

The analysis of triacylglycerols by high-temperature gas chromatography, along the last 10 years has been reviewed in this paper. The interest in this topic has grown along the last years due to the triacylglycerols are the main components of oils and fats and they are being used for the characterization and authentication of foods products. The most commonly used procedures, including the official methodologies, applying high-temperature gas chromatographic techniques are shown. Their importance in the characterization of different kind of samples, vegetable oils, seeds, dairy products, etc., is considered. This review is not intended to be a comprehensive dissertation on the field of triacylglycerols analysis since that would require sufficient space to occupy a book in its own right. Rather, it will outline selected considerations and developments, where the technique has been applied.

An IUPAC-based approach to estimate the detection limit in co-extraction-based optical sensors for anions with sigmoidal response calibration curves.

An approach based on IUPAC methodology to estimate the limit of detection of bulk optode-based analytical methods for anions has been developed. The traditional IUPAC methodology for calculating the detection limit was modified to be adapted to particular cases where the calibration curves have a sigmoidal profile. Starting from the different full theoretical models for every co-extraction mechanism of the analyte in the membrane in bulk optodes, several particular simplified models at low analyte concentration were obtained and validated. The slope of the calibration curve at low analyte concentration was calculated from the first derivative of the simplified equation and, subsequently, the detection limit was estimated. This fitted-for-purpose estimation strategy was applied to anion quantification for in-house bulk optode-based analytical methods, and the estimated limits of detection were compared with those obtained by applying classical geometrical methodology. This way of establishing the detection limit yields values that maintain their true statistical and probabilistic aspects. It can be easily applied to any analytical system which yields non-linear calibration curves at low analyte concentration.

Olive oil quantification of edible vegetable oil blends using triacylglycerols chromatographic fingerprints and chemometric tools.

The present work studies the effectiveness of the use of triacylglycerols (TAGs) for the quantification of olive oil in blends with vegetable oils. The determinations were obtained using high-performance liquid chromatography (HPLC) coupled to a Charged Aerosol Detector (CAD), in combination with Partial Least Squares (PLS) regression and using interval PLS (iPLS) for variable selection. Results revealed that PLS models can predict olive oil concentrations with reasonable errors. Variable selection through iPLS did not improve predictions significantly, but revealed the chemical information important in the chromatogram to quantify olive oil in vegetable oil blends.

Discriminating olive and non-olive oils using HPLC-CAD and chemometrics.

This work presents a method for an efficient differentiation of olive oil and several types of vegetable oils using chemometric tools. Triacylglycerides (TAGs) profiles of 126 samples of different categories and varieties of olive oils, and types of edible oils, including corn, sunflower, peanut, soybean, rapeseed, canola, seed, sesame, grape seed, and some mixed oils, have been analyzed. High-performance liquid chromatography coupled to a charged aerosol detector was used to characterize TAGs. The complete chromatograms were evaluated by PCA, PLS-DA, and MCR in combination with suitable preprocessing. The chromatographic data show two clusters; one for olive oil samples and another for the non-olive oils. Commercial oil blends are located between the groups, depending on the concentration of olive oil in the sample. As a result, a good classification among olive oils and non-olive oils and a chemical justification of such classification was achieved.

Application of selected ion monitoring to the analysis of triacylglycerols in olive oil by high temperature-gas chromatography/mass spectrometry.

The analysis of the triacylglycerol (TAG) composition of oils is a very challenging task, since the TAGs have very similar physico-chemical properties. In this work, a high temperature-gas chromatographic method coupled to electron ionization-mass spectrometry (HT-GC/EI-MS), in the Selected Ion Monitoring (SIM) mode, method was developed for the analysis of TAGs in the olive oil; this is a method suitable for routine analysis. This method was developed using commercially available standard TAGs. The TAGs studied were separated according to their equivalent carbon number and degree of unsaturation. The peak assignment was carried out by locating the characteristic fragment ions having the same retention time on the SIM profile such as [RCO+74](+) and [RCO+128](+) ions, due to the fatty acyl residues on sn-1, sn-2 and sn-3 positions of the TAG molecule and the [M-OCOR](+) ions corresponding to the acyl ions. The developed method was very useful to eliminate the interferences that appeared in the mass spectrum since electron ionization can prevent satisfactory interpretation of spectra.

Characterization of recovery profiles using gas chromatography-triple quadrupole mass spectrometry for the determination of pesticide residues in meat samples.

The assessment of the recovery factor with the analyte concentration in meat samples has been studied for the determination of organochlorine and organophosphorus pesticides in meat by gas chromatography-triple quadrupole mass spectrometry (GC-MS/MS). For that purpose, recent IUPAC recommendations, which distinguishes between two terms, recovery factor and apparent recovery, have been followed. Besides, the systematic error due to the matrix effect has been evaluated by a new term recently proposed, calibration recovery. Recovery profiles were obtained analyzing spiked blank matrix, where the analytes were added before and after the extraction procedure. In a first step, the quantification of the compounds was carried out using a solvent calibration curve. The systematic errors due to the matrix effect during the quantification step and the error due to the sample treatment have been evaluated. Both apparent and calibration recovery components depend on the actual analyte concentration in the sample while the recovery factor remains constant except for analyte concentration close to quantification limit. In addition, the concentration limits, from which an acceptable recovery value (70-110%) can be obtained, are given. If spiked samples are quantified by matrix-matched calibration, the matrix effect is minimized and the calibration recovery component is 100%, and apparent recovery only depends on the recovery factor. The obtained values indicate recovery factor does not depend on the analyte concentration, except for those values closed to quantification limit.

Setting up of recovery profiles: A tool to perform the compliance with recovery requirements for residue analysis.

The use of the recovery term has presented some confusion in Analytical Chemistry. Recent IUPAC recommendations propose to distinguish between two terms: recovery or recovery factor, Re, and apparent recovery, Re*. Apparent recovery includes recovery factor and a new recovery term proposed in this paper, named calibration recovery, Re(C), which depends of the type of systematic error due to the matrix effect (constant and/or proportional) and is related to the applied calibration methodology. This paper highlights the dependence of the calibration recovery on the sample analyte concentration and, for extension, of the apparent recovery, defines the recovery profile, and makes evident the need to determine a "fit for purpose" analyte concentration interval to comply with a regulated recovery requirements. An approach to estimate the calibration recovery and its associated uncertainty in relation to the above-mentioned dependence is presented. The usefulness of the proposed methodology has been shown in the quantification of a pesticide by GC-ECD for assessing dermal exposure.

Determination of albumin in biological fluids by flow injection analysis using the peroxyoxalate chemiluminescent system in micellar medium.

A new sensitive analytical procedure for the determination of albumin is presented, based on the participation of a fluorescent derivative of the protein in the peroxyoxalate chemiluminescent system using imidazole as a catalyst. The chemiluminescent emission is detected in a flow injection analysis (FIA) system, employing micellar medium as carrier, whereas the derivatisation reaction is carried out off-line using fluorescamine as a label. The optimisation of the operational and chemical variables of the method such as concentration of reagents, flow-rates, injection volume, etc., involves the use of experimental designs, including Draper-Lin small composite design, scarcely used in analytical chemistry. The performance characteristics were established and a detection limit of about 0.1 fmol of albumin was obtained. The method has been validated and satisfactorily applied to the determination of albumin in human serum and urine.

A comparative study of the correction of systematic errors in the quantitation of pyrethroids in vegetables using calibration curves prepared using standards in pure solvent.

A comparative study of two mathematical approaches was performed in order to correct systematic errors due to the presence of the unexpected interferences which appear when the quantitation of the analyte in real samples is carried out with calibration curves built using standards in pure solvent. These methods consisted in the establishment of different mathematical expressions which transform the concentration (Cs) obtained using calibration graphs built using pure solvent into the corrected concentration (C(M)) that should be obtained if the quantitation is carried out with calibration curves built using standards dissolved in blank matrix extracts. In the two approaches the correction is performed from the results of an intermediate precision study which was carried out using both calibration graphs (prepared using pure solvent and blank matrix extract). By using ANCOVA to compare the slope of both solvent-based and matrix-matched calibration graphs, matrix effect was found in the determination of deltamethrin in tomato and acrinathrin in tomato and pepper. In these cases, both approaches led to good results.

Applying non-parametric statistical methods to the classical measurements of inclusion complex binding constants.

A study on using non-parametric statistical methods was carried out to calculate the binding constant of an inclusion complex and to estimate its associated uncertainty. First, a correct evaluation of the stoichiometry was carried out in order to ensure an accurate determination of the binding constant. For this purpose, the modified Benesi-Hildelbrand method had been previously applied. Then, four statistical methods (three non-parametric methods: two bootstrap approaches, the jackknife method and a parametric one: Fieller's theorem) were employed in order to compute the binding constant. The results obtained from applying these methods and the combination of the methods: jackknife after bootstrap and bootstrap after jackknife were compared. The best results in terms of accuracy were obtained from the application of a bootstrap method: the resampling residuals approach. These procedures were applied to the inclusion complex 2-hydroxil-propyl-beta-cyclodextrin-2,4-dichloro-phenoxyacetic, which shows photochemically-induced fluorescence.

Simultaneous quantification of chlorophenoxyacid herbicides based on time-resolved photochemical derivatization to induce fluorescence in micellar medium.

In this paper a sensitive and simple method for the resolution of mixtures of chlorophenoxyacid herbicides using photochemical derivatization induced fluorescence has been described. These compounds do not show any fluorescence, hence photolysed to induce fluorescence after direct irradiation with ultraviolet light in presence of a cationic surfactant (cetyltrimethylammonium chloride). Critical variables such as the surfactant concentration and the irradiation time have been optimised for each compound using Sequential Response Surface Methodology (SRSM) by applying Doehlert designs in order to obtain maximum fluorescence intensity. The difference shown between the optimised irradiation times for the formation of the photoproducts allowed us to propose a time-resolved photoactivation method, for the simultaneous determination of binary mixtures, based on the use of different linear calibration curves established at various irradiation times depending on the mixture to be resolved. Satisfactory recoveries were obtained in the analysis of several mixtures of these herbicides at different ratios in spiked waters.

A new approach to qualitative analysis of organophosphorus pesticide residues in cucumber using a double gas chromatographic system: GC-pulsed-flame photometry and retention time locking GC-mass spectrometry.

A qualitative method for the screening of organophosphorus pesticides (OPs) that could present in different types of vegetables has been established and validated. A typical multi-residue extraction procedure of OPs using ethyl acetate and sodium sulphate has been applied. No clean-up was required after extraction, and concentrated extracts were analysed by gas chromatography with pulsed-flame photometric detection (GC-PFPD). Confirmation of compound identities was performed by gas chromatography with mass spectrometric detection (GC-MSD) in the electron impact (EI) mode with full scan acquisition. Retention time locking (RTL) software was used in order to improve the method capability of identification and confirmation. Spiked samples at pesticide concentrations equal to the maximum residue level (MRL) were used to check chromatographic performance and for validation studies. The proposed method allows a rapid and accurate identification of the studied OPs until the ng ml(-1) range for those whose use is forbidden, and above their MRL concentration for the rest.

Matrix-effects of vegetable commodities in electron-capture detection applied to pesticide multiresidue analysis.

The influence of the sample matrix in the analysis of pesticides in vegetable samples has been studied in order to determine if the matrix content introduces a systematic or proportional (or both) bias in the measurements. Experiments have been carried out during a 4-month period, in which calibration curves, prepared in solvent and in vegetable matrix, were prepared and analysed. A statistical treatment has been applied in order to: (i) check the stability of such calibrations during the period studied; (ii) compare both solvent and matrix-matched calibrations; and (iii) obtain a correction function. Applying the correction function to the results obtained with a solvent calibration it is possible to make a prediction of the values obtained applying a matrix-matched calibration. The performance of the correction function has been validated with recovery data. Finally the uncertainty derived from the use of each calibration plot and the correction function has been calculated.

Estimation and correction of matrix effects in gas chromatographic pesticide multiresidue analytical methods with a nitrogen-phosphorus detector.

The assessment of matrix effects in the quantification of organophosphorus pesticides in fruit and vegetables by GC-NPD, were studied applying ANCOVA. Calibration curves prepared in solvent were compared with calibration curves prepared in a blank matrix extract for eight different commodities, establishing whether the matrix induces systematic or proportional errors in the quantification of the pesticides. In such cases correction functions were obtained and validated by quantifying spiked samples using solvent calibrations and applying the correction functions to the data obtained. The results were compared with those obtained by quantification using matrix-matching calibrations and with those from 100% recovery experiments. It was found that the matrix effects can be avoided using the correction functions. Finally the contribution of the correction functions to the uncertainty of the results was estimated as well as their stability during a four month period.

Optimizing analytical methods using sequential response surface methodology. Application to the pararosaniline determination of formaldehyde.

Sequential response surface methodology is a general procedure to re-optimize common analytical methods on the basis of the application of the response surface methodology and of a new approach to the steepest ascent method. This procedure, which is easy to apply, consists of estimating an analytical function relating the response with the experimental parameters by means of a second-degree polynomial. Thus, a 2nd order design covering the total experimental domain is used and when a maximum is obtained, the characteristics of the response surface are confirmed using a new design, which is obtained contracting the first one. In the proposed methodology, Box-Behnken designs are used because they offer advantages in comparison with second order designs more frequently used in the steepest ascent method (central composite designs), i.e. fewer experiments are needed, they are more efficient, they can be moved through the experimental domain and they can even be easily contracted or expanded.

A first and straightforward approach to detection limits for narrow-bore HPLC and CE.

Two methodologies have been proposed for a simple statistical estimation of the detection limits in microbore-HPLC and CE techniques. Due to the impossibility to measure the blank, by using the extrapolated or approximated expressions it is possible to obtain a straightforward approach of the detection limits from the study of the residuals of the calibration data set, avoiding tedious treatment of the signals. The utility of these expressions has been checked on the data from an experimental microbore HPLC system.

On the estimate of capillary electrophoresis blanks: application to detection limits evaluation as recommended by IUPAC.

A new method to obtain the signal associated with an electrophoretic blank is presented to be included in the calibration procedures. The signal assigned to the blank is obtained by direct integration of the background noise using extrapolated values of the base-peak width at different concentrations in order to obtain the zero concentration. Detection limits that are better adjusted to a statistical evaluation are thus implemented, as recommended by the International Union of Pure and Applied Chemistry (IUPAC). The detection limit has been examined for a series of linear alkyl benzenesulfonates (C10-C13) and total linear alkyl benzenesulfonates that are widely used in many different commercial products.