Multi-Residue Analysis of 18 Regulated Mycotoxins by LC/MS/MS

Significance of the Topic

Mycotoxins are toxic metabolites produced by fungi that frequently contaminate cereals, nuts, milk and other agricultural commodities. Their diverse structures and potent health effects have led to strict global regulations, particularly within the European Union, where reporting limits are defined in EC 1886/2006. Reliable multi-residue analysis is essential to ensure food safety, compliance with legal thresholds and protection of public health.

Objectives and Scope

This study aimed to establish a single ultra high performance liquid chromatography tandem mass spectrometry method capable of quantifying 18 regulated mycotoxins in food and feed. The method was designed to achieve limits of quantification at or below EU maxima and to complete each injection cycle within 12.5 minutes.

Methodology

Samples were extracted by Scientific Analysis Laboratories following validated protocols and spiked with carbon-13 internal standards. Analysis employed multiple reaction monitoring transitions optimized for each compound. Calibration standards covered the EC-defined concentration ranges and allowed assessment of linearity and sensitivity.

Instrumentation

• UHPLC System: Shimadzu Nexera
• Column: Reversed phase, 100 mm × 2.1 mm, maintained at 40 °C
• Mobile Phase A: Water with additives; B: Methanol with additives; flow rate 0.4 mL/min
• Gradient: 15 % B at 0 min, ramp to 25 % at 1 min, 40 % at 2 min, 41 % at 4.5 min, 100 % at 7.5–10 min, re-equilibration to 15 % at 10.1 min
• Mass Spectrometer: Shimadzu LCMS-8060 triple quadrupole in ESI +/− mode with polarity switching
• Dwell times 10–40 ms, pause time 1 ms

Main Results and Discussion

The method achieved limits of quantification between 0.1 and 10 μg/kg for aflatoxins and 0.4 to 40 μg/kg for other compounds, matching EU requirements. Calibration curves exhibited coefficients of determination above 0.998 for all analytes. Chromatographic separation effectively resolved closely eluting isomers such as 3-acetyl-DON and 15-acetyl-DON within the 12.5-minute cycle. Representative MRM chromatograms confirmed high sensitivity and selectivity across diverse mycotoxin classes.

Benefits and Practical Applications

By consolidating multiple analytes into a single rapid method, laboratories can improve throughput and reduce solvent consumption. The robust performance supports routine monitoring in QA/QC and regulatory testing environments. Carbon-13 internal standards enhance quantitation accuracy in complex matrices.

Future Trends and Prospects

• Integration of high-resolution mass spectrometry for non-target screening and confirmation of emerging mycotoxins
• Automation of sample preparation workflows to increase throughput and reproducibility
• Expansion of multi-residue panels to include new fungal metabolites and co-contaminants
• Development of ambient ionization techniques for faster analysis without extensive chromatography

Conclusion

This work demonstrates a versatile UHPLC-MS/MS approach for the rapid and sensitive quantification of 18 regulated mycotoxins. With validated performance against EU limits and an efficient 12.5-minute cycle, the method offers a comprehensive solution for food safety laboratories.

Reference

1) A Rahmani, S Jinap, F Soleimany 2009 Qualitative and Quantitative Analysis of Mycotoxins Comprehensive Reviews in Food Science and Food Safety 8 202–251
2) M E Zain 2011 Impact of Mycotoxins on Humans and Animals Journal of Saudi Chemical Society 15(2) 129–144
3) M Sameni, A Dübecke, J F Weber 2014 Simultaneous Multi-Residue Determination of Mycotoxins in Foods Using LC-MS/MS Journal of Environmental & Analytical Toxicology 5(2) 1000259
4) J W Bennett, M Klich 2003 Mycotoxins Clinical Microbiology Reviews 16(3) 497–516
5) Mycotoxins regulations for Food accessed 6 September 2016