Comprehensive 2D GC with Dual Mass Spectrometry / Flame Ionization Detection for the Analysis of the Unsaponifiable Fraction of Vegetable Oils

Importance of the topic

Analysis of the unsaponifiable fraction of vegetable oils provides critical information on oil quality, traceability, and authenticity. While fatty acid profiles may overlap among different oils, the minor unsaponifiable components offer specific molecular fingerprints that can reveal adulteration and confirm origin.

Objectives and study overview

This work aimed to develop and apply a comprehensive two-dimensional gas chromatography (GC×GC) approach with dual flame ionization detection (FID) and mass spectrometry (MS) for both qualitative and quantitative characterization of the entire unsaponifiable fraction of vegetable oils without preliminary thin-layer chromatography (TLC) separation.

Methodology and instrumentation

Sample preparation involved saponification of 1 g oil with KOH/ethanol under reflux, extraction of the unsaponifiable fraction with diethyl ether, washing, drying over sodium sulfate, and evaporation. Derivatization to trimethylsilyl ethers was performed using BSTFA with TMCS in pyridine at 70 °C.

Instrumentation included a Shimadzu GC-2010 system equipped with an AOC-20i autosampler, Zoex dual-stage cryogenic modulator, and GCMS-QP2010 Ultra. GC×GC separations employed a SLB-5ms primary column (30 m×0.25 mm×0.25 µm) and an Rxi-17Sil secondary column (2 m×0.25 mm×0.25 µm). Helium was used as carrier gas in constant linear velocity mode. The first oven ramp was from 90 °C to 325 °C at 3 °C/min; the second from 140 °C to 360 °C (5 min) at 3 °C/min. Modulation period was 5 s. Dual uncoated transfer lines directed effluent to FID and MS branches. MS parameters: EI ionisation, 40–600 m/z scan at 20 000 amu/s, source 200 °C, interface 280 °C. FID operated at 360 °C with H₂, air, and helium makeup.

Main results and discussion

GC×GC–FID/MS produced well-resolved two-dimensional fingerprints of hydrocarbons, squalene, tocopherols, aliphatic alcohols, phytol, and sterols. In extra-virgin olive oil (EVOO), linear alkanes spanned C21–C33 with squalene dominant; only α-tocopherol was observed; aliphatic alcohols ranged C20–C30; sterol profiling revealed abundant cycloartenol, 24-methylene-cycloartenol, and β-sitosterol as key markers, alongside erythrodiol.

Sunflower oil showed lower squalene, both α- and β-tocopherols, alkanes from C23–C33, alcohols up to C32, and distinct sterol ergosta-7-en-3β-ol not present in EVOO. Some co-elution challenges in one-dimensional GC were resolved by GC×GC, improving quantitation of β-sitosterol and other sterols.

Quantitative analysis of eight EVOO samples complied with regulatory limits (e.g., cholesterol ≤0.5 %, brassicasterol ≤0.1 %, β-sitosterol ≥93 %). Comparison of one- and two-dimensional data indicated slightly lower apparent sterol contents in GC×GC due to enhanced resolution.

Benefits and practical applications

• Elimination of TLC speeds up sample preparation and reduces solvent use.
• Enhanced sensitivity and separation power allow clear class-pattern visualisation.
• Dual FID/MS detection delivers simultaneous quantitative and structural data.
• Robust fingerprinting supports authenticity testing and quality control in food analysis.

Future trends and applications

Integration of automated GC×GC workflows into routine QA/QC laboratories could streamline authenticity screening for diverse oils. Expanding spectral libraries and chemometric algorithms will enhance rapid classification. Emerging detectors and micro-modulation technologies may further boost sensitivity and throughput.

Conclusion

The developed GC×GC–FID/MS methodology offers a powerful means to comprehensively profile unsaponifiable fractions in vegetable oils. It outperforms one-dimensional GC by providing superior resolution, higher sensitivity, and detailed group-type patterns that are critical for reliable quality assessment and adulteration detection.

References

Technical Report C146-E255, Shimadzu Corporation, First Edition February 2015.