Multi-residue analysis of 18 regulated mycotoxins by LC-MS/MS

Importance of the Topic

Fusarium mycotoxins are structurally diverse fungal metabolites that commonly contaminate grains, spices and other foodstuffs. Their presence poses significant health risks to humans and animals, and regulatory authorities such as the European Union have established maximum levels to minimize exposure.

Objectives and Study Overview

This study set out to develop a single, rapid LC-MS/MS method for the simultaneous determination of 18 regulated mycotoxins – including aflatoxins B1, B2, G1, G2; fumonisins B1, B2, B3; ochratoxin A; and various trichothecenes – with limits of quantification at or below EU regulatory thresholds and an analysis cycle time of 12.5 minutes.

Methodology

Validated solvent extracts were prepared and spiked with 13C-labelled internal standards during extraction. Mobile phase additives (ammonium acetate, formate, fluoride and acetic acid) were evaluated. The optimal system used 0.15 mM ammonium fluoride (Solvent A) and 0.15 mM ammonium fluoride with 2% acetic acid in methanol (Solvent B) to enhance positive-ion signal intensity. Calibration covered relevant concentration ranges with correlation coefficients above 0.993.

Used Instrumentation

• Nexera UHPLC system
• LCMS-8060 triple quadrupole mass spectrometer
• Mastro PFP column (100×2.1 mm, 3 µm)
• 13C-labelled internal standards

Main Results and Discussion

• Ammonium fluoride mobile phase markedly increased positive-ion signal intensity compared to other additives.
• PFP column achieved near baseline resolution of regioisomer pairs 3-ADON/15-ADON and FB2/FB3, which coelute on C18 phases.
• Adding 2% acetic acid minimized fumonisin carryover without sacrificing sensitivity.
• Matrix tests in mixed spice and pepper extracts yielded repeatability with RSD below 10% for key toxins over 12 injections.

Benefits and Practical Applications

• High sensitivity and compliance with EU maximum limits.
• Short cycle time supports high-throughput routine analysis.
• Applicable to both PFP and C18 stationary phases and diverse food matrices.

Future Trends and Opportunities

Emerging developments may include expanded mycotoxin panels, novel gas-phase basic additives to further boost ionization, and integration of automated sample preparation workflows. Advances in stationary phase chemistries could improve isomer separation and analysis robustness.

Conclusion

The described LC-MS/MS method leverages ammonium fluoride mobile phases and a PFP column to deliver rapid, sensitive and robust multi-mycotoxin analysis. Its performance in complex food matrices demonstrates its suitability for routine quality control and regulatory compliance testing.

References

• Baker D.; Titman C.; Loftus N.; Horner J. Multi-residue analysis of 18 regulated mycotoxins by LC-MS/MS; Shimadzu Application Note PO-CON1767E; 2017
• European Commission. EC/1886/2006 Guidance on maximum levels for mycotoxins in food and feed