Multiresidue Pesticide Analysis in Onion by a Modified QuEChERS Extraction and Ion Trap GC–MSn Analysis

December 2, 2009
David Steiniger

Thermo Fisher Scientific, Inc.

,
Eric Phillips

Thermo Fisher Scientific, Austin, Texas

,
Jessie Butler

Thermo Fisher Scientific, Austin, Texas

The Application Notebook

The Application Notebook, The Application Notebook-12-02-2009, Volume 0, Issue 0

Most recently formulated pesticides are smaller in molecular weight and designed to break down rapidly in the environment.

David Steiniger, Jessie Butler and Eric Phillips, Thermo Fisher Scientific, Austin, Texas, USA.

Introduction

Most recently formulated pesticides are smaller in molecular weight and designed to break down rapidly in the environment.

The determination of pesticides in fruits and vegetables has been simplified by a new sample preparation method, QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe), published as AOAC Method 2007.01.

The sample preparation is simplified by using a single step buffered acetonitrile (MeCN) extraction and liquid–liquid partitioning from water in the sample by salting out with sodium acetate and magnesium sulphate (MgSO4). This note describes the application of the QuEChERS sample preparation procedure to analysis by gas chromatography–tandem mass spectrometry (GC–MSn ) on the Thermo Scientific ITQ 700 GC-ion trap mass spectrometer.

The study was performed to determine the linear ranges, quantification limits and detection limits for a long list of pesticides that are commonly used on onion crops, prepared in matrix using the QuEChERS sample preparation guidelines.

Once the calibration curve was constructed, multiple matrix spikes were analysed at levels of 100, 200 or 300 ng/g (ppb) and low-level spikes of 5, 10, 15, 25 or 50 ng/g (ppb) were used to verify the precision and accuracy of the analytical method.

Experimental Conditions

MSn parameters were optimized with the use of variable buffer gas, the testing of the isolation efficiency and adjustment of the collision induced dissociation (CID) voltage. A surge splitless injection was made into a 35% diphenyl/65% dimethyl polysiloxane column, (Thermo Scientific TRACE TR-35MS).

Sample extraction and clean-up

The QuEChERS sample prep procedure consists of three steps: the extraction, the clean-up and a solvent exchange. Extraction begins by adding 15 g of a thoroughly homogenized sample of onion into a 50 mL FEP extraction tube. 15 mL of 1% glacial acetic acid MeCN extraction solvent was poured into the tube on top of the sample. For the method validation and method detection limit samples, the surrogate and the pesticide solutions were spiked into this MeCN layer.

The tube was capped and vortexed for 30 seconds. The powder reagents were poured slowly into the MeCN layer. The 50 mL extraction tube was vortexed for 30 seconds until all of the powder reagents were mixed with the liquid layers. The tube was placed on a mechanical shaker for 5 minutes and then centrifuged for 5 minutes at 3000 rpm. Next, 11 mL of the top MeCN layer was removed and transferred to a 15 mL clean-up tube. This tube was capped and vortexed for 30 seconds and centrifuged for 5 minutes at 3000 rpm. A 5 mL aliquot of the top layer was transferred into a clean test tube for solvent exchange.

Solvent exchange

The 5 mL aliquot of extract was evaporated to dryness under a gentle stream of nitrogen at 40 °C. A 900 µL aliquot of hexane/acetone (9:1) was added and 100 µL of the internal standard, d10-parathion, was spiked into the organic solution. The tube was capped and vortexed for 15 seconds. The 1 mL of extract was transferred to a 2 mL clean-up tube, capped tightly and vortexed for 30 seconds. After centrifuging for 5 minutes at 3000 rpm, 200 µL of the clear extract was transferred to an autosampler vial with a small glass insert for injection on the ITQ 700. The individual calibration levels were spiked into each extract for the calibration curve in matrix before the final clean-up step.

Injection

The ITQ 700 is paired with the Thermo Scientific FOCUS GC gas chromatograph. The SSL inlet temperature was set to 250 °C. A 5 mm i.d. splitless liner selected for the surged pressure injection. For the surge splitless injection, the inlet pressure was held at an elevated pressure of 250 kPa for the 0.5 minute injection (splitless) time. This technique reduces the vapour cloud of a 2 µL injection from 0.37 mL to 0.19 mL. At an elevated injection flow rate of 4.6 mL/min, the liner was swept several times during injection. The target compounds had less time to interact with the inside walls of the liner, minimizing the breakdown of the more fragile pesticides.

A performance solution consisting of DFTPP, endrin and 4,4'-DDT was analysed as a daily check to determine system activity. The analysis of endrin and its breakdown products as part of daily quality control can alert the analyst that the system has developed active sites and maintenance is needed, decreasing the cost of running the analysis and saving time. The breakdown check results showed < 5% endrin breakdown on a daily basis.

Table 1

Separation

Chromatographic separation was achieved by using a 35% diphenyl/65% dimethyl polysiloxane column (30 m x 0.25 mm i.d., film thickness of 0.25 µm with a 5 m guard column). Interactions within the stationary phase showed a loss of certain pesticides at concentrations below 100 pg. A constant column flow rate of 1 mL/min was used for the duration of the run.

Detection

The detection of the pesticides was performed using the ITQ 700 ion trap mass spectrometer with optional MSn mode. This scanning mode offers enhanced selectivity over scanning modes such as full scan and selected ion monitoring (SIM).

The ITQ 700 operated in the MSn mode performs tandem MS functions by injecting ions into the ion trap and destabilizing matrix ions, isolating only the pesticide ion. CID energy is applied to fragment into its respective product ions. Finally these unique product ions are scanned out to generate the product ion spectrum. Because of the elimination of matrix interferences, this process produces more accurate results at the lower levels.

Results and Discussion

Linearity

The calibration curve was spiked into the onion matrix. Levels ranged from 1 ng/g to 1200 ng/g, depending on the compound and its MRL in onion. The linearity for most compounds was R2 > 0.995.

Limits of detection and Quantification

The actual limit of detection and limit of quantification were determined by preparing matrix spikes at a level near or below the MRL. Concentrations of 5, 10, 15, 25 and 50 ng/g were analysed in 10 matrix samples and the LOD and LOQ calculated from these results by multiplying the standard deviation by 2.821 and 10 respectively. The recovery of the 10 standards ranged from 79–159% with an average of 116%.

Method validation results

The method validation calculations were performed using Thermo Scientific EnviroLab Forms data analysis and reporting software on 12 matrix samples spiked at concentrations of 100, 200 and 300 ng/g. These samples had an average recovery of 104%, with an average % RSD of 22%.

Conclusions

The Thermo Scientific ITQ 700 GC-ion trap MS was showed excellent accuracy at low concentrations for a large number of pesticide residues analysed in onion. Using the instrument's MSn functionality allows the user to identify, confirm and quantify in one analytical run.

ITQ 700 is a registered trademark of Thermo Fisher Scientific.

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