Determination of 17 Mycotoxins in Cereals and Cereal Based Food Using Liquid Chromatography–Triple Quadrupole Mass Spectrometry

Importance of the Topic

The contamination of cereals by fungal metabolites known as mycotoxins represents a major food safety concern worldwide. Approximately one quarter of all crops may carry dangerous levels of these compounds, prompting strict regulatory limits in the EU. Effective multi-residue screening tools are essential for protecting consumer health and ensuring compliance in routine food and feed analysis.

Objectives and Study Overview

This study evaluates a streamlined dilute-and-shoot approach coupled with liquid chromatography–triple quadrupole mass spectrometry for the simultaneous determination of 17 mycotoxins. It addresses both the EU-regulated toxins (deoxynivalenol, zearalenone, T-2/HT-2 toxins, fumonisins, aflatoxins, ochratoxin A) and emerging compounds such as acetyl-deoxynivalenols, deoxynivalenol-3-glucoside, nivalenol, fusarenon X and sterigmatocystin. Key performance parameters—limits of detection/quantification, linearity, precision and matrix effects—were assessed in wheat and maize matrices.

Instrumentation Used

• Thermo Scientific UltiMate 3000 UHPLC system
• Thermo Scientific TSQ Endura triple quadrupole mass spectrometer
• Thermo Scientific Barnstead EASYpure II water purification system
• TraceFinder software (version 3.2) for instrument control and data processing

Methodology

Samples are homogenized, then 5–10 g portions are extracted with 80/20 acetonitrile–water for 60 min. After centrifugation, supernatants are diluted 1:5 with water, filtered through a 0.2 µm PTFE syringe filter and injected (10 µL) onto an Accucore C18 column (100 × 2.1 mm, 2.6 µm) at 40 °C. A binary mobile phase of water/methanol (98/2 to 2/98) containing 5 mM ammonium acetate and 1% acetic acid is used over a 12 min gradient. The TSQ Endura operates in timed-SRM with positive/negative polarity switching and optimized collision energies for each analyte.

Results and Discussion

Limits of detection and quantification achieved in solvent, wheat and maize ranged below or near EU maximum levels for all regulated mycotoxins. Chromatographic peaks were generally symmetrical, with nivalenol showing slight broadening. Matrix effects assessed by slope comparison indicated moderate to strong signal suppression in maize, justifying the use of matrix-matched calibration. Linearity exceeded R² > 0.99 for most analytes across relevant concentration ranges. Injection precision (RSD) remained below 20% in nearly all cases.

Benefits and Practical Applications

The dilute-and-shoot workflow minimizes sample preparation time and solvent use, while enabling comprehensive multi-mycotoxin quantitation. It meets EU regulatory requirements for cereals and cereal-based products and can be readily adopted for routine food quality control laboratories.

Future Trends and Opportunities

To address stricter limits in infant and baby foods, further sensitivity gains may be achieved via solid-phase extraction cleanup or more sensitive instruments (e.g., TSQ Quantiva). Incorporating isotopically labeled standards, automated sample handlers and high-throughput platforms will support enhanced accuracy, robustness and productivity in large-scale monitoring programs.

Conclusion

The presented LC-MS/MS method offers a rapid, robust and regulatory-compliant solution for simultaneous analysis of 17 mycotoxins in cereal matrices. With minor modifications for improved sensitivity, it is suitable for both general cereal testing and stringent baby food applications.

Reference

• Bennett et al. Clin Microbiol Rev. 2003;16:497–516.
• Marroquín-Cardona et al. Food Chem Toxicol. 2014;69:220–230.
• Commission Regulation (EC) No 1881/2006.
• Regulation (EC) No 1126/2007.
• Recommendation 2013/165/EU on T-2 and HT-2 toxins.
• Commission Regulation (EU) No 165/2010 amending aflatoxin limits.
• EFSA Mandate M-2013-0260 on acetylated deoxynivalenols.
• EFSA scientific opinion on sterigmatocystin. EFSA Journal. 2013;11(6):3254.