Analyze organophosphorus pesticides in the apple matrix by GC/MS/FPD using an Agilent J&W DB-35ms Ultra Inert GC Column

Organophosphorus pesticides are widely used in the agricultural industry for crop protection. Human toxicities for this class of molecules have shown acute and chronic effects from pesticide poisoning. OP pesticides affect the ner­vous system of insects and mammals by inhibiting an enzyme, acetylcholinesterase, that is important in helping regulate nerve impulses.1

Children are considered more susceptible to organophosphate toxicity because their pesticide dose per body weight is larger compared to that of adults.2 Children also have lower levels of detoxifying enzymes that deactivate OP pesticides, con­tributing to their vulnerability to pesticide exposure.3,4 Recent studies have shown a correlation between OP pesti­cide exposure and an increased risk for attention deficit hyperactivity disorder and other neurodevelopmental deficits in children.5-7 Because the main source of exposure for children is through consumption of food contain­ing OP pesticide residues, analytical testing capable of determining residual pesticides in food samples is critical.8

The multiresidue determination of pesticides in fruits and veg­etables usually involves an organic extraction of the pesti­cides from the plant matrix, followed by a cleanup procedure to remove co-extractives and other interferences. Anastassiades and colleagues developed the QuEChERS [quick, easy, cheap, effective, rugged, and safe] method for the analysis of pesticide residues in produce.9 This approach simplifies the traditional, labor-intensive extraction and cleanup procedure, while providing a fast, robust, and cost-effective method suitable for extracting pesticide residues.

Flow chart of the Agilent QuECHERS extraction procedure for apple samples

Figure 1: Flow chart of the Agilent QuECHERS extraction procedure for apple samples

Chromatographically active compounds such as OP pesticides can adsorb onto active sites in the sample flow path, particularly at trace levels, compromising an ana­lyte’s response. These pesticides tend to show peak tailing through interaction with active sites in a chromatographic system. This makes analysis challenging, particularly in diffi­cult sample matrices. Minimizing activity in the gas chromatography column is essential to ensure accurate quantitation. Agilent’s J&W DB-35ms Ultra Inert column minimizes column activity so difficult and active analytes can be consistently analyzed at trace levels. The use of the mid-polarity DB-35ms UI phase also offers additional selectivity over a non-polar phase, which can assist in resolving potentially co-eluting peaks, or shift a peak of interest away from matrix interferences.

Chromatographic Conditions

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Table 1: Chromatographic Conditions
Table 2: Flow Path Supplies
Table 3: Correlation Coefficients for the OP Pesticides Calibration Standards Analyzed by GC/MS-SIM and FPD in Phosphorus Mode with a Split Ratio for MSD:FPD = 3:1

A gas chromatographic system capable of multisignal detec­tion can provide complementary data for identification, confirmation, and quantitation of target analytes from a single injection. This method provides simultaneous detection of OP pesticides by gas chromatography/ mass spectrometry/selected ion monitoring and flame photometric detector in phosphorus mode by splitting the column effluent between the mass selective detector and FPD. The approach chosen here uses a GC/MSD/FPD system to identify and confirm the order of elu­tion for peaks of interest. Once the elution order is estab­lished, the chromatographic parameters can easily be trans­ferred to a GC/FPD system. The use of FPD detection without flow splitting is expected to increase sensitivity threefold, fur­ther improving lower level detection.

The GC/MS system was also equipped with backflush capa­bility, which shortens instrument cycle time by backflushing late-eluting matrix components through the inlet purge valve. Long bakeout times between injections are avoided by using this technique. Backflushing has the additional benefit of increasing the time intervals for source cleaning by effectively clearing deleterious matrix components from the system.10


An Agilent 7890A GC, combined with an Agilent 5975C GC/MSD equipped with a flame photometric detector and an Agilent 7683B auto­matic liquid sampler, were used for this series of experiments. A purged two-way capillary flow technology device was used to split the effluent 3:1 to the MSD:FPD. The CFT device also allowed for post-column backflush. Table 1 lists the chro­matographic conditions used for these analyses. Table 2 lists the flow path consumable supplies used in these experi­ments.

Reagents and Chemicals

All reagents and solvents were high-performance liquid chromatography or Ultra Resi grade. Acetonitrile from Honeywell (Muskegon, Mich.), toluene from Honeywell Burdick & Jackson, and acetone from JT Baker were purchased through VWR International (Radnor, Pa.). The 12-component custom pesticide standard was prepared by Ultra Scientific (N. Kingstown, R.I.).

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