Determination of PAH4 in edible oils with automated sample preparation

Significance of the Topic

The monitoring of polycyclic aromatic hydrocarbons (PAHs) in edible oils is critical due to their carcinogenic and mutagenic properties and the lipophilic accumulation in fats. European legislation (Regulation (EC) No 1881/2006 and No 835/2011) limits benzo[a]pyrene to 2 µg/kg and the sum of four PAHs (PAH4) to 10 µg/kg in oils, demanding highly sensitive, reliable analytical methods for routine quality control.

Objectives and Study Overview

This work aimed to transfer and modernize a published DACC–HPLC–FLD method onto the Thermo Scientific Vanquish Core platform, automating sample preparation via online solid phase extraction (SPE) and achieving a total analysis time of 1 hour. Key goals were to quantify benzo[a]pyrene, benzo[a]anthracene, chrysene, and benzo[b]fluoranthene (PAH4) with sensitivity at least ten times below the legal limits, and to simplify sample handling compared to traditional offline procedures.

Methodology and Instrumentation

Sample Preparation:

• Weigh 1 g oil into 5 mL flask, dilute with isopropanol, warm to 35 °C, and filter (0.45 µm PTFE).
• Inject 500 µL of filtrate.

Chromatographic Setup:

• Thermo Scientific Vanquish Core HPLC comprising System Base C, two Quaternary Pump Cs (loading and analytical), Split Sampler CT, Column Compartment C with two 2-position valves, and Fluorescence Detector F (dual PMT, 8 µL cell).
• Trap column: ChromSpher Pi DACC (3 × 80 mm, 5 µm); Analytical columns: two Nucleodur PAH (125 × 4 mm and 100 × 4 mm, 3 µm).
• Mobile phases: water (A), acetonitrile (B), isopropanol (C); dual-pump gradient with online backflush to remove lipids.
• FLD settings optimized by online 3D excitation/emission scans; multi-step wavelength program covering 270–290 nm excitation and 385–430 nm emission phases.

Main Results and Discussion

FLD Optimization and Interferences:

• Distinct excitation/emission settings yielded clear detection of each PAH; minor co-elution observed between perylene and benzo[b]fluoranthene but interference negligible (<3 %).
• Benzo[b]chrysene internal standard overlapped with matrix peaks in crude oils; calibration performed without internal standard using blank oil matrix.

Trap Efficiency and Calibration:

• Online trapping showed analyte recoveries within ±10 % compared to direct injection, confirming efficient retention and transfer from DACC column.
• Limit of quantification: 0.1 µg/kg for each PAH4 with signal-to-noise >19 (S/N >10 threshold).

Sample Analysis:

• Crude and virgin oils contained detectable PAH4 levels; synthetic triglyceride showed no PAHs.
• Crude coconut oil exhibited PAH4 concentrations up to ~90 µg/kg, exceeding calibration range.
• Method precision: RSD <5 % across replicate injections.
• Spike recoveries ranged from 91 % to 131 % depending on sample matrix.
• Carryover negligible after 500 µg/kg injection (<0.1 %).

Benefits and Practical Applications

This automated online SPE–HPLC–FLD approach reduces analysis time to 1 hour (vs. 80 min and 8–10 h for offline methods), minimizes solvent consumption and manual handling, and achieves sensitivities one order of magnitude below regulatory limits. The method is well suited for high-throughput quality control in food laboratories.

Future Trends and Potential Applications

Opportunities for advancement include coupling the DACC trap to mass spectrometric detection for broader PAH profiling, adopting improved sorbent chemistries to enhance selectivity, scaling up sample throughput via multiplexed SPE modules, and extending the workflow to other lipid-rich matrices such as dairy, meat, and environmental samples.

Conclusion

The transfer of an established DACC–HPLC–FLD method to the Vanquish Core platform successfully automates sample cleanup and analysis of the EU-regulated PAH4 in edible oils. The protocol delivers robust, sensitive, and reproducible results within a markedly reduced timeframe, fulfilling legislative and customer requirements.

References

1. Commission Regulation (EU) No 835/2011 amending Regulation (EC) No 1881/2006 on PAHs in food.
2. Alves da Silva S. et al., Food Chemistry, 221 (2017), 809–814.
3. Barranco A. et al., J. Chromatogr. A, 988 (2003), 33–40.
4. Barranco A. et al., Food Chemistry, 86 (2004), 465–474.
5. van Stijn F. et al., J. Chromatogr. A, 750 (1996), 263–273.
6. Jing C. et al., Thermo Scientific Application Note 196, 2016.
7. DIN EN ISO 22959:2009 – PAHs by on-line DACC and HPLC-FLD.
8. Franz H. and Jendreizik V., Fluorescence Method Development Handbook, 2013.