Coupling of Vac-HS-SPME and GC×GC-qMS for simultaneous 5-HMF quantification and volatile profiling in honey

Importance of the Topic

The formation of 5-hydroxymethylfurfural (5-HMF) in honey is a reliable indicator of thermal treatment and storage history. Regulatory limits under EU Directive 2001/110/EC ensure product freshness and quality, with maximum concentrations ranging from 15 to 80 mg/kg depending on origin and enzyme activity.

Objectives and Study Overview

This study aims to develop a rapid, sensitive, and green workflow for simultaneous quantification of 5-HMF and untargeted profiling of semi-volatile organic compounds (sVOCs) in honey. Integrating vacuum-enhanced headspace solid-phase microextraction (Vac-HS-SPME) with comprehensive two-dimensional gas chromatography coupled to quadrupole mass spectrometry (GC×GC-qMS) provides improved throughput compared to conventional HPLC-UV or UV spectrophotometric methods.

Methodology and Instrumentation

The optimized protocol uses 0.5 g of decrystallized honey in a 20 mL vial. After quick freeze and one-minute vacuum application, Vac-HS-SPME extraction occurs at 60 °C for 17 minutes under vacuum. On-fiber derivatization with 50 µL BSTFA + 1% TMCS at 80 °C for five minutes follows. Thermal desorption at 270 °C with cryogenic focusing at 0 °C precedes GC×GC separation. The first dimension employs an apolar column, the second a polar column, and detection uses qMS in combined SIM and full-scan mode. Fiber type: DVB/CAR/PDMS, 1 cm.

Main Results and Discussion

Method validation demonstrated linear response for 5-HMF from 5 to 80 µg/g, with LOD of 1.6 µg/g and LOQ of 4.7 µg/g. Matrix-matched calibration showed an average recovery of 98% and precision of 21% RSD. Analysis of six commercial samples yielded an average repeatability of 8% RSD and a 6% bias compared to HPLC-UV references. Comprehensive GC×GC-qMS profiling revealed key sVOCs such as benzaldehyde, nonanal, menthofuran, and phenylethanol, aiding authenticity assessment.

Benefits and Practical Applications

• High sensitivity for 5-HMF quantification well below regulatory limits.
• Broad chemical coverage of sVOCs for origin and freshness markers.
• Automatable, reduced analysis time versus official protocols.
• Reduced sample size and solvent use align with green analytical chemistry.

Future Trends and Potential Applications

Advances may include coupling to high-resolution MS for non-target screening, miniaturized vacuum modules for on-site testing, and integration with chemometric models for authenticity and geographical origin verification.

Conclusion

The Vac-HS-SPME/GC×GC-qMS approach provides a robust, green, and comprehensive solution for simultaneous 5-HMF quantification and volatile profiling in honey. It outperforms traditional methods in speed, sensitivity, and environmental footprint, supporting quality control and authenticity studies.

References

1. Eggermont D., Pardi F., Purcaro G. Coupling of Vac-HS-SPME and GC×GC-qMS for simultaneous 5-HMF quantification and volatile profiling in honey.
2. E. Psillakis, Anal. Chim. Acta 2017.
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5. International Honey Commission. Harmonised Methods, 2009.