Fully Automated Mycotoxin Analysis in the ppq Range FREESTYLE ThermELUTE™

Importance of the Topic

The accurate and sensitive detection of mycotoxins in food matrices at ultra-trace levels is critical for consumer safety and regulatory compliance. Automated sample preparation combined with high-performance chromatography streamlines laboratory workflows, increases throughput, reduces human error, and enables reliable quantification in the parts-per-quadrillion (ppq) range.

Objectives and Study Overview

This study demonstrates a fully automated workflow for mycotoxin analysis—specifically Aflatoxin M1 and B/G—in milk and related products. The goal is to validate the FREESTYLE ThermELUTE™ robotic platform coupled with miniaturized immunoaffinity SMART columns and HPLC detection, achieving ultra-low detection limits and high reproducibility.

Methodology and Instrumentation

Sample Preparation and Cleanup

• Extraction and dilution follow SMART column protocols, including dilution in phosphate-buffered saline.
• Automated processing via FREESTYLE ThermELUTE™:
• Loading: up to 20 mL extract (1 mL/min).
• Washing: 4 mL (1 mL/min).
• Thermal elution: 98 °C for 8 minutes to dissociate toxin-antibody complexes.
• Eluate directly transferred to the HPLC sample loop.

Instrumental Setup

• FREESTYLE ThermELUTE™ robotic system for full automation.
• AflaCLEAN M1 SMART and AflaCLEAN SMART immunoaffinity columns (3 cm length).
• HPLC with Dionex 3400 RS detector.

Main Results and Discussion

Sensitivity and Detection Limits

• Aflatoxin M1 was quantified down to 500 ppq (0.5 ppt) in milk, demonstrating linear responses at 10 ppt, 1 ppt, 0.9 ppt, 0.8 ppt, and 0.5 ppt levels.
• Aflatoxin B/G analysis achieved detection of individual congeners (B1, B2, G1, G2) at total toxin loads as low as 0.0025 ng/10 mL (0.9–3.6 ppt range).

Reproducibility

• Triplicate analyses of independent spiked samples showed consistent retention times and peak areas, indicating high method precision even at ultra-trace concentrations.

Sensitivity Validation

• Comparison of blind and spiked samples confirmed clear discrimination and accurate quantification at 500 ppq for Aflatoxin M1.

Benefits and Practical Applications

• Throughput: up to 500 samples per week with minimal hands-on time.
• Cost Reduction: miniaturized SMART columns consume less solvent and reduce waste.
• Robustness: thermal elution ensures complete recovery of bound toxins without organic solvents.
• Versatility: compatible with any HPLC system, allowing seamless integration into existing laboratory infrastructures.

Future Trends and Applications

Advancements in automation and column miniaturization are expected to further lower detection limits and expand application to other toxin classes and complex food matrices. Integration with mass spectrometry detectors and smart data analytics will enhance specificity, quantitation accuracy, and real-time monitoring capabilities.

Conclusion

The FREESTYLE ThermELUTE™ platform, combined with SMART immunoaffinity columns and HPLC detection, provides a fully automated, sensitive, and reproducible method for mycotoxin analysis at the ppq level. This workflow meets stringent regulatory requirements while boosting laboratory efficiency and reducing operational costs.

Reference

LCTech GmbH. Fully Automated Mycotoxin Analysis in the ppq Range with FREESTYLE ThermELUTE™, Application Note AS012, November 2019.