Determination of aflatoxins in dried fruit samples - from extraction to high efficient detection

Significance of the Topic

Aflatoxins are toxic secondary metabolites produced by Aspergillus species that frequently contaminate dried fruits under warm and humid conditions. Their ingestion poses serious health risks, including carcinogenic effects. Reliable monitoring methods are essential to ensure consumer safety and compliance with international regulations.

Objectives and Study Overview

This study aims to develop a streamlined extraction protocol and a highly sensitive HPLC method with photochemical post column derivatization for simultaneous detection of aflatoxins B1 B2 G1 and G2 in dried fruit samples. The goal is to achieve detection limits well below regulatory thresholds while minimizing matrix interferences.

Methodology and Instrumentation

Sample preparation involves three sequential steps

• Solid liquid extraction with methanol water to solubilize aflatoxins and remove solids
• Liquid liquid extraction with n hexane and chloroform to eliminate fats and hydrophobic matrix compounds
• Solid phase extraction on silica cartridges to purify analytes and reduce residual interferences

Chromatographic analysis employs a C18 reverse phase column at 60C coupled to a fluorescence detector with excitation at 365 nm and emission at 460 nm. Photochemical post column derivatization at 254 nm enhances fluorescence of B1 and G1 without using toxic halogen reagents. The HPLC system includes a binary pump autosampler thermostat interface box and dedicated software for data acquisition.

Main Results and Discussion

The optimized cleanup resulted in chromatograms with minimal matrix peaks in the critical elution window. Limits of detection were 0.05 ng per mL for aflatoxins B1 and G1 and 0.015 ng per mL for B2 and G2, surpassing European Commission requirements of 2 and 4 ppb. Recovery experiments in spiked matrices including mixed dried fruits peanuts pistachio and cereal purees demonstrated high reproducibility and robustness.

Benefits and Practical Applications

This approach enables simultaneous quantification of all major aflatoxins in a single run with high sensitivity and repeatability. It is well suited for quality control laboratories food safety monitoring and regulatory compliance testing across diverse food and feed matrices.

Future Trends and Applications

Advances in UHPLC technology and integrated photochemical derivatization promise further reductions in analysis time and detection limits. Expanding the protocol to multi mycotoxin workflows and coupling with mass spectrometry could broaden the scope of food safety surveillance.

Conclusion

The combined extraction and HPLC fluorescence method with photochemical derivatization provides a reliable sensitive and reproducible platform for aflatoxin analysis in dried fruits. It meets stringent regulatory standards and offers versatility for routine food safety applications.

Used Instrumentation

• AZURA P6.1L binary pump
• AZURA AS 6.1L autosampler
• RF 20A fluorescence detector
• UVE photochemical derivatization module
• Eurospher II C18 column 150 x 4.6 mm ID
• AZURA CT 2.1 thermostat
• IFU 2.1 LAN interface
• ClarityChrom 8.1 software

References

1. Hedayati MT Pasqualotto AC Bowyer P Denning DW Aspergillus flavus human pathogen allergen and mycotoxin producer Microbiology 153 1677 1692 2007
2. World Health Organization Aflatoxins Food safety digest Department of food safety and zoonoses 2018
3. US FDA Action levels for aflatoxins in animal food CPG Sec 683.100 2019
4. European Commission Regulation 401 2006 methods of sampling and analysis for mycotoxins in foodstuffs 2016
5. European Commission Regulation 1881 2006 setting maximum levels for contaminants in foodstuffs 2006
6. Folmert K Margraf M Monks K Quick and easy determination of aflatoxins in food matrices with photochemical post column derivatization AppNote VFD0178 2019