gc-ms/ms analysis and data acquisition for assessing pesticides in avocado samples prepared by the quechers method

Análisis de plaguicidas en aguacate utilizando el método QuEChERS y CG-MS/MS

In chemical analysis, the matrix effect (ME) can be defined in various ways, but a widely accepted general definition is as follows: it refers to the change in the signal, particularly a change in the slope of the calibration curve when the sample matrix or portion of it is present during the analysis of a specific analyte. As a critical aspect, ME necessitates thorough investigation during the validation process of any analytical method, as it directly affects the accuracy of quantitative measurement for the target analytes1. Ideally, a sample pretreatment procedure should be selective enough to avoid extracting any components from the sample matrix. However, despite significant efforts, many of these matrix components still end up in the final determination systems in most cases. Consequently, such matrix components often compromise the recovery and precision values, introduce additional noise, and escalate the overall cost and labor involved in the method.
In gas chromatography (GC), ME arises due to the presence of active sites within the GC system, which interact with the target analytes through various mechanisms. On the one hand, the matrix constituents block or mask these active sites that would otherwise interact with the target analytes, resulting in frequent signal enhancement2. On the other hand, active sites that remain unobstructed may cause peak tailing or analyte decomposition due to strong interactions, leading to a negative ME. However, this can offer potential benefits in certain cases2. It is crucial to emphasize that achieving complete inertness in a GC system is exceedingly challenging, despite using highly inert components and proper maintenance. With continuous use, the accumulation of matrix components in the GC system becomes more pronounced, causing an increased ME. Nowadays, it is widely recognized that analytes containing oxygen, nitrogen, phosphorus, sulfur, and similar elements, exhibit a greater ME as they readily interact with these active sites. Conversely, highly stable compounds such as hydrocarbons or organohalogens do not undergo such interactions and do not show observable ME during analysis2,3.
Overall, ME cannot be fully eliminated, leading to the development of several strategies for compensation or correction when complete removal of matrix components is not feasible. Among these strategies, the utilization of deuterated internal standards (ISs), analyte protectants, matrix-matched calibration, the standard addition method, or the modification of injection techniques have been documented in scientific literature1,2,4,5. The SANTE/11312/2021 guidelines have also recommended these strategies6.
Regarding the application of matrix-matched calibration to compensate for MEs, sample sequences in practical situations encompass diverse types of foods or various samples from the same commodity. In this case, the assumption is made that employing any sample from the same commodity will effectively compensate for ME in all samples. However, there is a lack of sufficient studies in the existing literature that specifically investigate this issue7.
The multiresidue determination of pesticides in matrices containing an appreciable percentage of fat and pigments constitutes a challenging task. The considerable amount of coextracted material can significantly affect the extraction efficiency and interfere with the subsequent chromatographic determination, potentially damaging the column, source, and detector, and resulting in significant MEs8,9,10. Consequently, the analysis of pesticides at trace levels in such matrices necessitates a significant reduction of matrix components before analysis while ensuring high recovery values7. Obtaining high recovery values is crucial to ensure that pesticide analyses remain reliable, accurate, and compliant with regulatory standards. This is vital for ensuring food safety, environmental protection, and informed decision-making in agriculture and related fields.
Avocado is a fruit of high commercial value cultivated in tropical and Mediterranean climates worldwide and widely consumed both in its regions of origin and in the numerous export markets. From the analytical point of view, avocado is a complex matrix containing a significant number of fatty acids (i.e., oleic, palmitic, and linoleic), similar to nuts, a significant pigment content, as in green leaves, as well as sugars and organic acids, similar to those found in other fruits11. Due to its fatty nature, special attention must be given when employing any analytical method for analysis. While pesticide residue analysis has been conducted on avocados using GC-MS in some instances8,12,13,14,15,16,17,18,19,20, it has been relatively less frequent compared to other matrices. In most cases, a version of the Quick-Easy-Cheap-Effective-Rugged-Safe (QuEChERS) method has been applied8,12,13,14,15,16,17,18. None of these studies have investigated the consistency of MEs among different avocado varieties.
Therefore, the aim of this work was to study the consistency of MEs and recovery values for 45 representative pesticides across different varieties of avocado (i.e., Criollo, Hass, and Lorena) using the QuEChERS method with ammonium formate and GC-MS/MS. To the best of our knowledge, this is the first time that this type of study has been conducted on such fatty matrix samples.

Preparación de muestras de aguacate utilizando el método QuEChERS con formiato de amonio para el análisis de pesticidas

Determinación de 45 plaguicidas en variedades de aguacate por el método QuEChERS y cromatografía de gases-espectrometría de masas en tándem

To begin the sample preparation using the QuEChERS method, weigh five grams of an avocado sample of interest in a 50 milliliter centrifuge tube. Add 50 microliters of the procedural internal standard or P-IS solution to the tube. Add 10 milliliters of aceto nitrile to the centrifuge tube and shake the tube at 70 RPM for five minutes.Then add 2.5 grams of ammonium formate to it and shake the tube again at 70 RPM for five minutes. Subsequently, centrifuge the tube at 1, 800 G for five minutes. Next, in a two milliliter centrifuge tube, take the appropriate amounts of the reagents displayed on the screen.Then add one milliliter of the extract into the tube for purification using dispersive solid phase extraction. Vortex the tube for 30 seconds before centrifuging it at 1, 800 G for five minutes. Transfer 200 microliters of the purified extract to an auto-sampler vial, and add 20 microliters of the analyte protectant solution, followed by adding 50 microliters of a pre-prepared triphenyl phosphate solution.Finally, perform instrumental analysis using a GC-MS/MS system.

Análisis GC-MS/MS y adquisición de datos para la evaluación de plaguicidas en muestras de aguacate preparados por el método QuEChERS

Realice los análisis utilizando un sistema GC-MS/MS con un cuadrupolo triple equipado con una interfaz de ionización de electrones de 70 electronvoltios en un muestreador automático. Asegúrese de que las presiones de gas sean correctas. Helio a 140 PSI y argón a 65 PSI.Encienda el interruptor GC principal ubicado en el panel frontal. Y encienda el interruptor MS ubicado en la parte posterior. Abra el software de análisis en tiempo real GCMS que controla todos los parámetros del sistema GC-MS/MS.Haga clic en control de vacío. Luego avanzó seguido de una bomba rotativa para iniciar el sistema de vacío. Haga clic en iniciar para encender las bombas turbomoleculares uno y dos.A continuación, haga clic en la opción de inicio del calentador de fuente de iones. Ajuste la temperatura de la interfaz MS a 250 grados Celsius y la temperatura de la fuente de iones a 300 grados Celsius. Mantenga el horno GC a una temperatura inicial de 50 grados centígrados durante un minuto.Luego aumente a 180 grados Celsius a una velocidad de 25 grados Celsius por minuto. Posteriormente, aumente la temperatura a 230 grados centígrados a cinco grados centígrados por minuto, y luego a 290 grados centígrados a 25 grados centígrados por minuto. Luego mantenga la temperatura constante a 290 grados centígrados durante seis minutos.El tiempo total de análisis es de 24,6 minutos. Una vez que todos estos sistemas estén encendidos, haga clic en cerrar. A continuación, haga clic en la opción de ajuste del software de análisis y seleccione la vista del monitor de picos para realizar una verificación inicial de las condiciones del espectrómetro de masas.Haga clic en Adquisición y, en la ventana que se muestra, haga clic en Descargar parámetros iniciales. Verifique que el equipo esté listo para GC y listo para MS. Para la adquisición de datos, haga clic en nuevo archivo por lotes del software y cree una secuencia que contenga información como el nombre de la muestra, el ID de la muestra, el archivo de método, el archivo de datos, el volumen de inyección y el archivo de ajuste. Agregue filas según sea necesario y haga clic en guardar.Haga clic en Inicio por lotes y deje que comience el proceso de inyección. Realizar la inyección a 250 grados centígrados en el modo de lista dividida, manteniendo un volumen de inyección de un microlitro. Después de un minuto después de la inyección, abra el split.Analice los analitos utilizando el modo de monitorización de reacción múltiple, o MRM, que es el modo estándar empleado en los sistemas MS/MS con un cuadrupolo triple. Abra el software de análisis posterior a la ejecución para el análisis de datos. Haga clic en la inyección a analizar.Navegación a través de la tabla que contiene los analitos y selección del pico de interés. Haga clic en el pico o en la región de interés para visualizar el cromatograma. Revise las integraciones máximas y, si es necesario, realice la integración manual.Verificar las áreas de todos los analitos para realizar los cálculos necesarios y la evaluación del método. La generación de curvas de calibración emparejadas con matrices reveló un efecto de matriz relativamente consistente de menos del 20% clasificado como un ME suave para la mayoría de las combinaciones de plaguicidas. Los resultados de la figura mostrada demostraron el recuento de plaguicidas cuyos porcentajes promedio de recuperación estuvieron dentro del rango aceptable de 70 a 120% para cada tipo de aguacate.Una proporción significativa de los plaguicidas analizados exhibieron porcentajes de recuperación que se encontraban dentro de este rango específico, con valores relativos de desviación estándar por debajo del 20%

Research

JoVE Journal

Chemistry

GC-MS/MS Analysis and Data Acquisition for Assessing Pesticides in Avocado Samples Prepared by the QuEChERS Method

Determination of 45 Pesticides in Avocado Varieties by the QuEChERS Method and Gas Chromatography-Tandem Mass Spectrometry

Gas chromatography (GC) tandem mass spectrometry (MS/MS) stands as a preeminent analytical instrument extensively employed for the surveillance of ...

Investigación

Química