EU Priority PAH Analysis in Pumpkin Seed Oil Using Bond Elut EMR—Lipid Cleanup by GC/MS/MS

Significance of the Topic

Polycyclic aromatic hydrocarbons (PAHs) are lipophilic contaminants formed during high-temperature processing and environmental exposure. Certain PAHs, including benzo(a)pyrene and related compounds, are regulated in edible oils due to their carcinogenic potential. Pumpkin seed oil, produced by roasting and pressing seeds without refining, presents a complex lipid matrix prone to PAH contamination. Reliable, low-level quantitation in such fatty samples is critical for food safety and compliance with EU regulations.

Objectives and Overview of the Study

This study demonstrates a robust analytical workflow to quantify the 16 EU priority PAHs in pumpkin seed oil. The aim was to validate a combined liquid–liquid extraction and dispersive solid-phase cleanup approach using Bond Elut EMR—Lipid and PSA/C18/MgSO4, followed by gas chromatography–tandem mass spectrometry (GC/MS/MS) with enhanced source cleaning and backflush capabilities. Key performance metrics—recovery, precision, accuracy, linearity, and limits of quantitation—were evaluated against EU regulatory thresholds.

Methodology and Instrumentation

Samples (1 g oil) were extracted with acetonitrile, spiked with isotopically labeled PAH internal standards, and vortexed. After centrifugation, the supernatant underwent EMR—Lipid dSPE cleanup (with limited water to preserve PAH solubility), followed by EMR-Polish for water removal and a secondary PSA/C18/MgSO4 step to eliminate residual lipids and moisture. Extracts were concentrated tenfold and reconstituted in isooctane for analysis.

• GC/MS/MS System: Agilent 7890B GC with 7010 triple quadrupole, JetClean self-cleaning ion source, and Capillary Flow Technology backflush.
• Autosampler: Agilent 7693A.
• Column Configuration: Agilent Select PAH capillary column (5 m transfer segment for backflush, 25 m analytical segment).
• Operating Conditions: Inlet, transfer line, and source at 320 °C; hydrogen flow to source (0.33 mL/min); backflush after run to purge high-boiling residues.

Main Results and Discussion

The dual cleanup strategy provided superior lipid removal compared to single SPE methods, yielding cleaner baselines and extended column life. Calibration curves (1–100 ng/g, 1/x2 weighting) were linear (R2>0.99). Limits of quantitation were 1 ng/g for most PAHs; two high-molecular-weight compounds required 10 ng/g. Recovery ranged 50–120% for mid-eluting PAHs at 1, 10, and 50 ng/g; heavier PAHs met criteria only at 50 ng/g. Precision (RSD) was <20%, except benzo(a)pyrene at 1 ng/g (23%). Accuracy fell within 100±20% across levels. Backflush and JetClean minimized matrix deposition and maintained sensitivity for low-level PAHs.

Benefits and Practical Applications of the Method

This protocol offers a streamlined, reproducible workflow for routine quantitation of regulated PAHs in fatty matrices. It balances effective lipid removal with high analyte recovery, ensures instrument robustness, and meets stringent EU performance criteria. Laboratories performing QA/QC, regulatory monitoring, or research in food safety can implement this approach to enhance throughput and data confidence.

Future Trends and Potential Applications

Advances in matrix removal sorbents and automation may further lower LOQs and simplify workflows. Coupling enhanced cleanup with high-resolution mass spectrometry could improve selectivity for trace contaminants. Extending this approach to other edible oils, complex food matrices, and environmental samples will support broader contaminant surveillance. Ongoing development of miniaturized and in-line cleanup systems may enable real-time monitoring during processing.

Conclusion

The combined EMR—Lipid and PSA/C18/MgSO4 cleanup, followed by GC/MS/MS with JetClean and backflush, delivers a validated method for EU priority PAH analysis in pumpkin seed oil. The method achieves regulatory compliance for sensitivity, accuracy, and precision while protecting instrument performance in a challenging lipid matrix.

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