Correction of predicted concentration in the use of solvent-based calibration lines for determining carbendazim, fuberidazole and thiabendazole in water after a SPE step

This paper addresses an attempt to overcome the deviation that results from the use of a solid phase extraction (SPE) procedure for extracting trace levels of three benzimidazole pesticides (carbendazim, fuberidazole and thiabendazole) from water samples, for their subsequent quantitative determination by spectrofluorimetry, using univariate calibration. The deviation is due to an attenuation effect originating in the C(18) cartridge used in the SPE step. The approach developed is based on the calculation of a correction factor (fc) that is dependent on the signal measured after the SPE step. In order to calculate fc a study of the intermediate precision of two calibration graphs (with and without SPE) was performed. The fc was added to the predicted concentrations for the analytes using a calibration graph for pure solvent, built every time that the analysis is done. In addition, predictions were made using both average calibration graphs obtained from the intermediate precision study. In this study, the first of these three options was shown to improve the accuracy of predictions in the presence of matrix effects.     

Selection of a Representative Matrix for Calibration in Multianalyte Determination of Pesticides in Vegetables by Liquid Chromatography-Electrospray Tandem Mass Spectrometry

An analytical method for determining residues of twenty pesticides by liquid chromatography (LC) coupled to electrospray ionisation (ESI) tandem mass spectrometry (MS-MS) in eight commodities, cucumber, tomato, pepper, green bean, eggplant, zucchini, melon and watermelon, has been developed and validated. On one hand, calibration curves prepared in solvent were compared with calibration curves prepared in a blank matrix extract of each target matrix. On the other hand, calibration curves and recoveries for each commodity were compared. Cucumber was selected as potential reference matrix for the target vegetables.     

Multi-residue determination of 130 multiclass pesticides in fruits and vegetables by gas chromatography coupled to triple quadrupole tandem mass spectrometry

A multi-residue method has been developed and validated for the simultaneous quantification and confirmation of around 130 multiclass pesticides in orange, nectarine and spinach samples by GC-MS/MS with a triple quadrupole analyzer. Compounds have been selected from different chemical families including insecticides, herbicides, fungicides and acaricides. Three isotopically labeled standards have been used as surrogates in order to improve accurate quantitation. Samples were extracted by using accelerated solvent extraction (ASE) with ethyl acetate. In the case of spinach, an additional clean-up step by gel permeation chromatography was applied. Determination was performed by GC-MS/MS in electron ionization mode acquiring two MS/MS transitions for each analyte. The intensity ratio between quantitation transition (Q) and identification transition (q) was used as confirmatory parameter (Q/q ratio). Accuracy and precision were evaluated by means of recovery experiments in orange, nectarine, and spinach samples spiked at two concentration levels (0.01 and 0.05 mg/kg). Recoveries were, in most cases, between 70% and 120% and RSD were below 20%. The limits of quantification objective for which the method was satisfactorily validated in the three samples matrices were for most pesticides 0.01 mg/kg. Matrix effects over the GC-MS/MS determination were tested by comparison of reference standards in pure solvent with matrix-matched standards of each matrix. Data obtained showed enhancement of signal for the majority of analytes in the three matrices investigated. Consequently, in order to reduce the systematic error due to this effect, quantification was performed using matrix-matched standard calibration curves. The matrix effect study was extended to other food matrices such as raisin, paprika, cabbage, pear, rice, legume, and gherkin, showing in all cases a similar signal enhancement effect.     

Quantitative estimation of circulating metabolites without synthetic standards by ultra-high-performance liquid chromatography/high resolution accurate mass spectrometry in combination with UV correction

An early assessment of metabolite exposure in preclinical species can provide quantitative estimation on possible active or toxic metabolites. Frequently, synthetic metabolite standards are not available at the preclinical stage, precluding the quantitation of metabolites by means of calibration curves and quality control (QC) samples. We present here an approach to determine the extent of circulating metabolites using 'metabolite standards' generated by in vitro incubations in combination with the correction for mass spectrometry response based on UV response. The study was done by coupling ultra-high-performance liquid chromatography (UHPLC) to LTQ-Orbitrap high-resolution mass spectrometry, and the quantitation was based on full scan high-resolution accurate mass analysis in combination with retention time. First, we investigated the separation capacity of a 10.5 min UHPLC method and the quantitative capability of an LTQ-Orbitrap for full scan accurate mass quantitation by spiking chemical standards of buspirone and its six metabolites in blank plasma. Then we demonstrated the use of a UV correction approach to quantitatively estimate buspirone and its metabolites in plasma samples from a rat pharmacokinetics study. We compared the concentration versus time profiles of buspirone and its six metabolites in rat plasma samples obtained using three different approaches, including using UV correction, using individual standard curves for each metabolite prepared from the synthetic standard, and using a calibration curve of the parent compound buspirone. We demonstrated the estimated metabolite exposure of buspirone using this UV correction approach resulted in rank ordering of metabolite exposure within three-fold of the value obtained with metabolite standards, in contrast to eight-fold without UV correction. The approach presented in this paper provides a practical solution to an unmet bioanalytical need for quantitative information on metabolites without standards in preclinical in vivo studies.     

Effect of sample matrix on the determination of total petroleum hydrocarbons (TPH) in soil by gas chromatography–flame ionization detection

Research papers in different fields of analytics indicate that the effect of matrix-induced chromatographic response enhancement (matrix effect) is a commonly encountered problem in gas chromatography applications. In this paper, an example of the effect of sample matrix on the quantitative determination of total petroleum hydrocarbons (TPH) by GC–FID in soil is presented. Two types of soil were selected for the evaluation. Extraction and analysis of the soil samples was in accordance with CEN prEN 14039. The relative systematic error resulting from the matrix effect was obtained for three different TPH concentrations by statistical comparison of the slopes of the matrix-matched calibration lines and a pure solvent calibration line. Too high TPH concentrations were obtained when conventional solvent calibration was used for quantitation. This demonstrates that matrix-matched calibration should be exploited in the determination of petroleum hydrocarbons in soil samples. However, there was also significant enhancement of the response due to an interfering matrix with decreasing analyte concentration. Enhancement seems to be especially evident in the quantification of TPH over the concentration range encountered in polluted environments. As a result, even when matrix-matched calibration is used for quantitation, it is still necessary to establish the range over which a linear response can be expected. Otherwise too high results for sample TPH concentrations will be obtained.     

Rapid method for determination of chlormequat residues in tomato products by ion-exchange liquid chromatography/electrospray tandem mass spectrometry

A rapid method has been devised for the direct determination of chlormequat in tomato samples. No clean-up is required, and analysis uses ion-exchange liquid chromatography/tandem mass spectrometry interfaced with electrospray ionization (LC/ESI-MS/MS). A cation-exchange column was used with an aqueous ammonium acetate/acetonitrile mixture as the mobile phase under isocratic conditions. The method was validated in terms of detection limits (LOD), quantitation limits (LOQ), linearity, recovery, precision and accuracy. Good results in the low micro g kg(-1) level were obtained for the LOD and LOQ of chlormequat in tomato samples. Comparison of solvent and matrix-matched calibration curves demonstrated the absence of significant matrix effects and the feasibility of using external calibration. Linearity was established over two orders of magnitude by performing homoscedasticity and Mandel fitting statistical tests. The absence of both constant and proportional systematic errors was verified by evaluating the recovery function, demonstrating good method accuracy. Excellent precision in terms of intra-day repeatability was calculated (RSD% <3.4). Extraction recoveries from tomato products were calculated, by using a labelled internal standard (d(4)-chlormequat), to be in the 93 +/- 5-99 +/- 7% range. The applicability of the method to the determination of chlormequat residues in tomato products was demonstrated.     

Fast gas chromatography for pesticide residues analysis using analyte protectants

Fast GC-MS with narrow-bore columns combined with effective sample preparation technique (QuEChERS method) was used for evaluation of various calibration approaches in pesticide residues analysis. In order to compare the performance of analyte protectants (APs) with matrix-matched standards calibration curves of selected pesticides were searched in terms of linearity of responses, repeatability of measurements and reached limit of quantifications utilizing the following calibration standards in the concentration range 1-500 ng mL(-1)(the equivalent sample concentration 1-500 microg kg(-1)): in neat solvent (acetonitrile) with/without addition of APs, matrix-matched standards with/without addition of APs. For APs results are in a good agreement with matrix-matched standards. To evaluate errors of determination of concentration synthetic samples at concentration level of pesticides 50 ng mL(-1) (50 microg kg(-1)) were analyzed and quantified using the above given standards. For less troublesome pesticides very good estimation of concentration was obtained utilizing APs, while for more troublesome pesticides such as methidathion, malathion, phosalone and deltamethrin significant overestimation reaching up to 80% occurred. According to presented results APs can be advantegously used for "easy" pesticides determination. For "difficult" pesticides an alternative calibration approach is required for samples potentially violating MRLs. An example of real sample measurement is shown. In this paper also the use of internal standards (triphenylphosphate (TPP) and heptachlor (HEPT)) for peak areas normalization is discussed in terms of repeatability of measurements and quantitative data obtained. TPP normalization provided slightly better results than the use of absolute peak areas measurements on the contrary to HEPT.     

Application of multiple headspace-solid-phase microextraction followed by gas chromatography–mass spectrometry to quantitative analysis of tomato aroma components

The objective of this paper is to investigate the potential of multiple headspace-solid-phase microextraction (MHS-SPME) for the determination of volatile compounds in complex matrix samples. A method based on MHS-SPME for the determination of around 20 volatile compounds, responsible of tomato flavour and aroma has been developed and validated, using gas chromatography with mass spectrometry (ion trap analyser) for analysis. For this purpose, the experimental β parameter, resulting from the MHS-SPME theoretical development, has been obtained from real sample analysis (in triplicate) for each identified compound, carrying out up to 5 consecutive extractions. Later, this parameter is used to perform quantitation of real samples after just a single HS-SPME extraction. Precision, expressed as repeatability, has been evaluated by analysing six replicates of a real sample, showing relative standard deviations between 4 and 20%. For accuracy study, quantitative results have been compared with those obtained by means of standard additions on replicate samples, and no statistically significant differences between the two methods were observed. Since MHS-SPME uses the estimated total area corresponding to the complete extraction of compounds (obtained from the β parameter), quantitation can be carried out by external calibration using standards in solvent and splitless injection, instead of by SPME. Linearity, tested in the range 0.05–15 μg/mL, showed satisfactory values, with coefficients of correlation between 0.995 and 0.999. Limits of detection were in the range of 0.25–5 ng/g. MHS-SPME has been proved to be an adequate technique to avoid matrix effects in complex samples quantitation. Its applicability to the determination of volatile tomato components, together with its limitations, is discussed in this article.     

Capillary zone electrophoresis of orotic acid in urine with on-line isotachophoresis sample pretreatment and diode array detection

Two hydroxide-selective microbore analytical columns (the Dionex AS11 and AS15) were tested and compared for the quantitation of anionic species in 30% hydrogen peroxide. The ions of interest were fluoride, acetate, formate, chloride, bromide, nitrate, sulfate, and phosphate. Statistically sound calibration and spiking studies were carried out, investigating the range of a blank to 60 ppb. Prior to injection onto the separators, peroxides were loaded without pretreatment onto a concentrator column, which was then washed with deionized water to remove the matrix. Although retention times gradually decreased during the spiking studies, reliable quantitation was still achievable on both columns at the target concentration of 30 ppb. However, various resolution problems meant that the AS11 should not be recommended for this application.     

Application of matrix solid-phase dispersion to the determination of a new generation of fungicides in fruits and vegetables

A method based on matrix solid-phase dispersion (MSPD) and gas chromatography to determine eight fungicides in fruits and vegetables is described. Fungicide residues were identified and quantified using nitrogen-phosphorus detection and electron-capture detection connected in parallel and confirmed by mass spectrometric detection. The method required 0.5 g of sample, C18 bonded silica as dispersant sorbent, silica as clean-up sorbent and ethyl acetate as eluting solvent. Recoveries from spiked orange, apple, tomato, artichoke, carrot and courgette samples ranged from 62 to 102% and relative standard deviations were less than 15% in the concentration range 0.05-10 mg kg(-1). Detection and quantitation limits ranged 3-30 microg kg(-1) and 10-100 microg kg(-1), respectively, with linear calibration curves up to 10 mg kg(-1). The analytical characteristics of MSPD compared very favourably with the results of a classical multiresidue method, which uses ethyl acetate and anhydrous sodium sulphate for the extraction.     

Evaluation of evaporative light-scattering detector for combinatorial library quantitation by reversed phase HPLC

A quantitation study using reversed phase HPLC with UV and evaporative light-scattering detector (ELSD) was conducted on 90 library standards selected from 15 small molecule combinatorial libraries (six standards from each library). This study assessed the quantitation errors using a single calibration curve for rapid purity analysis of combinatorial libraries. The average quantitation error of six standards from one library at 200 microM by UV was 13. 4%, 20.6%, and 60.3%, at 214, 220, and 254 nm, respectively. By ELSD, the average quantitation error of these six standards at 200 micro was only 7.7%. Applying this ELSD calibration curve to 84 standards from 14 structurally diverse libraries, an average quantitation error of 16.4% was obtained. The average quantitation error of all 90 standards from 15 libraries using 15 calibration curves was 18.5%.     

Creatinine determination in urine samples by batchwise kinetic procedure and flow injection analysis using the Jaffé reaction: chemometric study

The classical Jaffé reaction for the determination of creatinine in urine samples is tested. A comparative study of the main analytical characteristics focussed to minimize the bias error and improve the precision, for the batchwise and flow injection (FI) methods is realized. Also, the effect of the albumin concentration in the determination of creatinine has been studied. Different analytical signals were studied. Absorbance increments at different times permit to estimate the creatinine concentration free from bias error in urine by the batchwise method using the calibration graph obtained with creatinine standards and no measurement of the blank solution is needed. The lineal interval was 0.92–50 mg l⁻¹ and seven samples can be processed per hour by an operator. No previous treatment of the urine sample is necessary. The FI method provides also good results. The lineal interval was 30–100 mg l⁻¹ and the sample rate was around 20 samples per hour. If increased albumin levels are detected in the urine, standard addition method or the calibration graphs with standards in presence of albumin are needed in order to obtain accurate results when FI method is employed. The obtained accuracy of the both methods allows its application as diagnostic tool to establish the urinary creatinine levels.     

Matrix-induced effects: a critical point in the gas chromatographic analysis of pesticide residues

The influence of several experimental factors related to the enhanced gas chromatographic responses yielding apparent recoveries of pesticide residues greater than 100% was investigated. Optimisation of a gel permeation chromatographic clean-up step with respect to the trueness and precision of generated data was performed. An increase of relative detector response (100%=response of analyte in pure solvent solution) was evidenced to be dependent both on the concentration of the analyte and the character of the matrix: pronounced matrix-induced effects were observed particularly in orange and wheat extracts at low concentration levels of analytes (especially for GC–electron-capture detection analysis of certain pesticides). As soon as the splitless injector became contaminated after injection of large series of matrix-containing samples, a decrease of relative responses of pesticides, largely below 100%, was experienced. Although troublesome compounds tending to give matrix-induced effects can be identified, and increased recoveries may be tentatively predicted, poor accuracy of generated data can be presumed as long as quantitation is not based on a standard prepared in blank matrix extract to compensate for matrix-induced effects.     

Data handling in HPLC-diode array UV-spectrometry

Summary The mathematical resolution of overlapped chromatographic peaks obtained in HPLC with a diode array UV detector has received considerable attention recently. The purpose of the proposed methods is the quantitation and identification of unknown solutes in complex mixtures by an efficient use of all available data. Basically two approaches have been proposed: the first one resolves the concentration profiles and calculates the pure spectra by applying a minimal number of assumptions, which is denoted self-modeling-curve resolution. The second one is based on a match of pure spectra available in a spectral library with the measured mixture spectra. In this paper both approaches are evaluated with respect to their performance to provide the pure spectra and an accurate quantitation of the concentrations.     

Determination of Surfactants Using Ion Pair Extraction of Near-infrared Laser Dyes

Abstract

Ionic surfactants and near-infrared laser dyes formed complexes which were extracted into organic solvents as ion pairs. Surfactants were determined spectrophotometrically by measuring the near-infrared absorbance and fluorescence of the ion pair in the organic solvent. Several of the commercially available near infrared dyes have been found suitable for surfactant determination in water using this technique. The excess near-infrared dye coextracted into the organic solvent was determined by blank extractions. The calibration curves were linear within two orders of magnitude of surfactant concentrations. Non-linear calibration curves are obtained for wider concentration range of surfactants. This method using the recently developed near-infrared laser diode intracavity technique was applied to the determination of SDS in water. Lower detection limits and ease of operation are the major advantages of using this new laser diode technique. The extraction efficiency of different solvent systems was evaluated.     

Correction function on biased results due to matrix effects: Application to the routine analysis of pesticide residues

A new strategy to carry out the correction of analytical results affected by systematic errors due to the matrix effect is proposed. Two types of external calibrations must be established with the purpose to estimate the matrix effect: solvent calibration (SC) and matrix-matched calibration (MC). These calibration curves are statistically compared and a correction function (CF) is proposed with the aim to simplify the resolution to the problems associated with the incidence of matrix systematic error in the analytical results. Applying this correction function to the results obtained from the solvent calibration, it is possible to make a prediction of the values that would be obtained when the matrix-matched calibration is applied. On the other hand, a rigorous study of the associated uncertainty is developed and applied to the calculated correction function. Finally, this correction function is validated by means of obtained data of recovery studies carried out by a traditional methodology. The methodology has been satisfactorily applied to the quantification of the pesticide procymidone by HPLC for assessing dermal exposure.     

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.     

Pesticide analysis in rose wines by micellar electrokinetic chromatography

In this work, the determination of 11 pesticides (pirimicarb, metalaxyl, pyrimethanil, procymidone, nuarimol, azoxystrobin, tebufenozide, fenarimol, benalaxyl, penconazole, and tetradifon) in rose wines by micellar EKC (MEKC) using reversed electrode polarity stacking mode (REPSM) as online preconcentration strategy is described. The MEKC buffer consisted of 100 mM sodium tetraborate and 30 mM SDS at pH 8.5 with 6% v/v 1-propanol. A solid-phase microextraction (SPME) procedure using PDMS/divinylbenzene (PDMS/DVB) fibers was applied to extract the selected pesticides from the rose wine samples. The comparison between the calibration curves obtained from hydroalcoholic solutions (12% v/v ethanol) and from rose wines (matrix matched calibration) showed the existence of a strong matrix effect. Furthermore, a comparison with calibration curves obtained with white wine samples also showed significant differences for most of the analyzed pesticides. As a result, a matrix matched calibration was developed. Quantitative extraction from spiked wine samples was carried out in triplicate at two levels of concentration (range 0.18-6.00 mg/L). LODs between 0.040 and 0.929 mg/L were achieved, which are below the maximum residue limits (MRLs) established for wine grapes (except for pirimicarb) by the EU and Spain legislation as well as by the Codex Alimentarius. The established method - which is solvent free, cost effective, and fast - was also applied to the analysis of several homemade rose wine samples and a commercial one. Two of the selected pesticides were found in some of the analyzed samples.     

Validated HPAEC-PAD method for prebiotics determination in synbiotic fermented milks during shelf life

Interest concerning functional food has been growing in recent years, and much attention has been focused on the choice of prebiotic fibers and probiotic microorganisms added to food products with the aim of improving health, producing synbiotic products. In the work reported here, an innovative analytical method performed by high-performance anion-exchange chromatography (HPAEC) with pulsed electrochemical detection has been optimized and validated for application to the study of prebiotic effects in synbiotic fermented milk prepared by addition of probiotics. The proposed method permits the evaluation of fructooligosaccharides and inulooligosaccharides with degrees of polymerization of 6–7 and 4–7, respectively. Quantitative determination was performed on oligosaccharides, whose standards are not commercially available, by employing calibration curves built by adding a known amount of the fiber used as an ingredient to the matrix. The work provides results from a parallel study on simultaneous variations of prebiotics and probiotics during the shelf life of fermented milk samples. The main advantage over time-consuming, classic enzymatic methods, whose results are limited only to average fiber content, is the possibility of dosing each carbohydrate by performing a single HPAEC run. Validation in terms of detection and quantitation limits, linearity, precision, and recovery was carried out.