DETERMINATION OF AFLATOXINS IN A WIDE RANGE OF FOOD AND AGRICULTURAL COMMODITIES USING IMMUNOAFFINITY CHROMATOGRAPHY COLUMN CLEAN-UP WITH UPLC OR HPLC WITH FLUORESCENCE DETECTION

Significance of the Topic

Aflatoxins are potent carcinogenic mycotoxins produced by Aspergillus species that contaminate a wide range of food and feed commodities worldwide. Chronic exposure poses serious health risks and can lead to regulatory enforcement actions, economic loss, and trade rejections. Sensitive, selective, and robust analytical methods are essential to ensure food safety and compliance with stringent global limits.

Objectives and Study Overview

This study evaluated the performance of an immunoaffinity clean-up using the new AflaTest WB SR+ column coupled with either HPLC or UPLC with fluorescence detection. The method was applied to seven matrices including corn, wheat, oats, chili powder, black pepper, cocoa, and a traditional Chinese medicine (Fallopia multiflora).

Methods and Instrumentation

A multistep procedure combined sample extraction, immunoaffinity column clean-up, and chromatographic analysis:

• Sample extraction: homogenization of 10–25 g aliquots with acetonitrile/water (90/10 or 80/20, v/v) followed by filtration.
• Immunoaffinity clean-up: AflaTest WB SR+ columns bind aflatoxins B1, B2, G1, G2, M1, M2, and sterigmatocystin. Columns were washed with water and eluted with acetonitrile/methanol (1:2, v/v).
• Derivatization and detection:
  • HPLC: Waters Alliance e2695 system with photochemical reactor and fluorescence detection (Ex360 nm/Em440 nm) on a Nova-Pak C18 column, isocratic water:methanol (55:45) at 0.8 mL/min, 50 µL injection.
  • UPLC: ACQUITY UPLC H-Class with fluorescence detector and large volume flow cell on an HSS T3 1.8 µm, 2.1×100 mm column, isocratic water:methanol (55:45) at 0.3 mL/min, 3–6 µL injection.

Main Results and Discussion

The method demonstrated excellent recoveries between 92 % and 116 % across all matrices with relative standard deviations below 8 %. UPLC provided baseline separation of all aflatoxins within seven minutes, and the large volume flow cell in UPLC-FLD eliminated the need for chemical derivatization. Limits of detection and quantification met or exceeded global regulatory requirements. No significant interferences were observed in blank samples.

Benefits and Practical Applications

• High sensitivity and selectivity for B- and G-aflatoxins and sterigmatocystin without additional derivatization reagents.
• Wide dynamic range (0.05–500 ng total aflatoxin), suitable for low-level infant foods and high-level feed monitoring.
• Versatility across diverse commodities including challenging matrices such as spices, cocoa, and botanicals.
• Reduced analysis time and simplified workflow with UPLC options.

Future Trends and Opportunities

Analytical advancements may include multiplexed immunoaffinity devices, integration with mass spectrometry for confirmatory analysis, greener solvent systems, fully automated workflows, and portable platforms for on-site screening. Continued development of high-resolution columns and detector technologies will further enhance throughput and robustness.

Conclusion

The described immunoaffinity-based clean-up coupled with HPLC or UPLC-FLD offers a comprehensive, reliable, and rapid solution for determining regulated aflatoxins in a wide array of food and agricultural commodities. The method meets AOAC performance criteria and supports routine compliance monitoring.

References

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• AOAC. Official Methods of Analysis. Appendix F Guidelines for Standard Method Performance Requirements, 2016.