Polycyclic Aromatic Hydrocarbon (PAH) Analysis in Fish by GC/MS Using Agilent Bond Elut QuEChERS dSPE Sample Preparation and a High Efficiency DB-5ms Ultra Inert GC Column

Significance of the Topic

The accurate determination of polycyclic aromatic hydrocarbons (PAHs) in seafood is critical for food safety and environmental monitoring. PAHs are persistent environmental contaminants with known carcinogenic potential that bioaccumulate in fish lipids following oil spills and other petrochemical releases. Rapid, reliable screening methods enable regulatory agencies and laboratories to assess contamination levels, protect public health, and support fishery reopening decisions.

Objectives and Study Overview

This study aimed to develop a streamlined analytical protocol for quantifying 16 priority PAHs in fish tissue at low to trace levels. The goals were to replace time-consuming multi-step cleanup procedures with a QuEChERS-based approach, integrate dispersive solid-phase extraction (dSPE) for matrix removal, and implement automated backflush in gas chromatography/mass spectrometry (GC/MS) to reduce cycle times and carryover.

Instrumentation

• Gas chromatograph: Agilent 7890 GC
• Mass spectrometer: Agilent 5975B MSD with selective ion monitoring (SIM)
• Inlet: Multimode inlet (MMI), splitless mode
• Autosampler: Agilent 7693 automatic liquid sampler (5 µL syringe)
• Column: Agilent J&W DB-5ms Ultra Inert, 20 m × 0.18 mm i.d., 0.18 µm film thickness
• Backflush device: Pressure controlled tee (PCT) post-column

Methodology

Sample preparation utilized a modified QuEChERS extraction with dSPE:

• Weigh 3 g of minced fish into a 50 mL tube and add surrogate/internal standards.
• Add 12 mL deionized water and 15 mL acetonitrile with ceramic mixing bars.
• Introduce QuEChERS salt packet (MgSO₄ and NaCl), vortex 1 min, centrifuge at 4000 rpm for 5 min.
• Transfer 8 mL acetonitrile layer to a dSPE tube containing cleanup sorbents, vortex and centrifuge.
• Analyze 1 µL aliquots by GC/MS in SIM mode, calibrating at 10, 25, 50, 100, 250, 500, and 1000 ng/mL.

Chromatographic conditions featured a temperature program from 50 °C to 315 °C, constant helium flow (1.7 mL/min), and post-run backflush at 315 °C for 7 min to remove late-eluting matrix components.

Main Results and Discussion

All 16 target PAHs were baseline-resolved in under 20 minutes. Calibration exhibited excellent linearity (R² 0.9988–0.9999). The method limit of quantitation was 10 ng/mL for benzo[a]pyrene, well below the 30 ng/g regulatory concern level for finfish. Recoveries at 25, 250, and 500 ng/mL spiking ranged from 80% to 139% with relative standard deviations below 6%, demonstrating high accuracy and precision. The automated backflush feature minimized carryover and eliminated lengthy bake-out cycles.

Benefits and Practical Applications

• Significant reduction in sample preparation time: up to 60 samples processed in an eight-hour shift versus 12–14 hours per sample by traditional methods.
• Robust cleanup of fatty fish matrices while maintaining low-level analyte sensitivity.
• Accelerated GC/MS cycle times and extended maintenance intervals through automated backflush.
• Suitable for routine screening to inform seafood safety and regulatory compliance.

Future Trends and Potential Applications

• Further miniaturization and automation of QuEChERS workflows.
• Integration of high-resolution MS for enhanced selectivity and non-target screening.
• On-line sample preparation coupling to GC/MS or LC/MS for continuous monitoring.
• Expansion to other complex food and environmental matrices.
• Development of portable or field-deployable PAH detection systems.

Conclusion

The described QuEChERS-dSPE sample preparation combined with a high-efficiency DB-5ms Ultra Inert GC column and automated backflush GC/MS offers a rapid, reliable, and high-throughput solution for trace PAH analysis in fish. The method meets regulatory performance criteria, significantly reduces labor and instrument downtime, and is adaptable for wider food safety and environmental applications.

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