Screening and Verifying Mycotoxins in Food with Q-TOF LC/MS and an Accurate Mass Library

Significance of the Topic

The contamination of food commodities by mycotoxins presents serious health concerns and strict regulatory demands. Broad-spectrum, high-resolution screening methods are essential to detect both regulated and emerging fungal metabolites in diverse matrices. Accurate mass LC/Q-TOF workflows can enhance sensitivity, selectivity, and retrospective data analysis capabilities for comprehensive mycotoxin surveillance.

Objectives and Study Overview

This application note reports on the development of an accurate-mass spectral library comprising over 300 mycotoxins and related metabolites. The library was integrated with two acquisition strategies—targeted MS/MS and All Ions MS/MS—using an Agilent 1290 Infinity LC coupled to an Agilent 6550 iFunnel Q-TOF. Forty-four representative mycotoxins were spiked into maize, hazelnut, and wine matrices to evaluate screening, identification, and verification performance.

Methodology and Instrumentation

Sample Preparation:

• Grind and homogenize 5 g of blank maize or hazelnut, add 20 mL acidified acetonitrile–water (79:20.9:0.1, v/v/v).
• Shake 90 min, settle solids, transfer supernatant, spike at three concentration levels.

Chromatographic Conditions:

• Column: Poroshell 120 EC-C18, 2.1×100 mm, 2.7 µm.
• Mobile phase A: 5 mM ammonium formate + 0.1% formic acid; B: same in methanol.
• Gradient: 10% B to 98% B over 10 min, total run 17 min, flow 0.40 mL/min, 30 °C, injection 2 µL.

Acquisition Modes:

• Target MS/MS: precursor selection, collision energy 20 V, 3 Hz scans for MS and MS/MS.
• All Ions MS/MS: simultaneous low-energy channel and two high-energy channels (0, 10, 40 V), 3 Hz.

Used Instrumentation

• Agilent 1290 Infinity UHPLC (Binary Pump, Autosampler, Column Compartment)
• Agilent 6550 iFunnel Q-TOF LC/MS
• Dual-spray Agilent Jet Stream electrospray source
• Agilent 1260 Isocratic Pump for reference mass delivery
• Agilent MassHunter Data Acquisition, Qualitative Analysis, and Quantitative Analysis software
• Custom Personal Compound Database and Library (Mycotoxins and Related Metabolites PCDL)

Main Results and Discussion

Both workflows effectively detected and confirmed 44 indicator mycotoxins in spiked matrices at 30 ng/mL. Mass deviations were generally below 1 ppm, with target scores above 90. Coelution scoring in All Ions MS/MS allowed verification of fragment ions without precursor isolation. Target MS/MS provided high-confidence spectra for low-mass analytes. In a naturally contaminated hazelnut sample, six mycotoxins were quantified, demonstrating applicability to real-world samples.

Benefits and Practical Applications

The approach offers rapid, minimal-prep screening and verification of regulated and emerging mycotoxins across complex matrices. It supports retrospective analysis for unknowns, high-throughput workflows, and complements single-analyte methods in QA/QC laboratories and research environments.

Future Trends and Applications

Expansion of accurate mass libraries to include newly discovered fungal metabolites, integration with automated data-mining tools, and application of ion mobility separation are expected to enhance selectivity. Harmonized, high-throughput HRAM workflows will facilitate regulatory monitoring and early detection of emerging mycotoxin threats.

Conclusion

The combination of accurate-mass spectral libraries with target and All Ions MS/MS acquisition on a high-resolution Q-TOF platform provides a robust strategy for comprehensive mycotoxin screening and verification in food. It yields high confidence identifications, broad compound coverage, and the potential for retrospective data review.

References

• Zöllner P.; Mayer-Helm B. J. Chromatogr. A. 2006, 1136, 123–169.
• Commission Regulation (EC) No. 1881/2006 and amendments for food contaminants.
• Commission Recommendation 2013/165/EU on T-2 and HT-2 toxins in cereals.
• Varga E. et al. Application Note, Agilent Technologies, 5991-4991EN, 2014.
• Varga E. et al. Application Note, Agilent Technologies, 5991-2808EN, 2013.
• Kempe G. et al. Application Note, Agilent Technologies, 5991-2227EN, 2013.
• Wüst B. et al. Application Note, Agilent Technologies, 5991-2295EN, 2013.