How Appropriate Clean-Up Can Improve the Robustness of an LC-MS/MS Method for the Determination of Multiple Mycotoxins in a Range of Food Matrices

Significance of the Topic

Mycotoxins are toxic secondary metabolites produced by fungi that commonly contaminate agricultural commodities. Their co-occurrence in food and feed poses serious health risks and economic challenges, driving demand for robust multi-toxin analytical methods. In high‐throughput testing laboratories, methods must combine sensitivity, speed, and reliability to support regulatory compliance and protect consumer safety.

Objectives and Study Overview

This study evaluates the long-term robustness of two multi-mycotoxin LC-MS/MS approaches—one employing a simple dilute-and-shoot protocol and the other incorporating a pass-through SPE clean-up step—over continuous operation. The goal is to determine which workflow better maintains analytical performance, reduces matrix interferences, and minimizes maintenance requirements.

Methodology and Instrumentation

• Sample preparation: Homogenized wheat samples spiked with 14 regulated mycotoxins. Two protocols compared: direct dilution (D&S) versus Oasis PRiME HLB pass-through SPE clean-up.
• LC-MS/MS system: ACQUITY UPLC I-Class PLUS coupled to a Xevo TQ-S micro, 8 µL injection volume, continuous runs 24 h/day for 35 days (~2 900 injections).
• Data evaluation: Control charts, linear regression to derive intermediate-precision slopes and t₁₀ (time to 10 % signal loss); matrix effect calculations; SCAN mode assessment of co-extractives; dried residue measurements; visual extract clarity.

Main Results and Discussion

• Intermediate precision: SPE reduced signal decline slopes by 45–89 % compared to D&S, increasing median t₁₀ from 22 to 68 days.
• Matrix effects: SPE clean-up lowered ion suppression/enhancement for most mycotoxins (up to 325 % reduction for fumonisins), improving quantification accuracy.
• Phospholipid removal: SPE virtually eliminated late-eluting phospholipid peaks, reducing background and charged surface build-up in the mass spectrometer.
• Co-extractives and maintenance: Total ion current in SCAN mode decreased by 6–18 % after SPE; dried residue mass decreased by 16–66 % across matrices; extracts appeared clearer post clean-up.

Benefits and Practical Applications

Implementing a pass-through SPE step enhances method robustness by reducing matrix interferences and charging effects that cause instrument drift. This leads to fewer maintenance interventions, lower failure rates, and more consistent quantification, ultimately boosting laboratory throughput and cost-effectiveness.

Future Trends and Opportunities

Advances may include automated inline clean-up, integration of isotopically labeled internal standards for further matrix compensation, expansion to emerging mycotoxins and complex matrices, and coupling with high-resolution mass spectrometry for broader screening.

Conclusion

The addition of an effective SPE clean-up to multi-mycotoxin LC-MS/MS workflows significantly prolongs stable instrument performance, reduces matrix effects, and minimizes system downtime. Laboratories adopting this strategy can achieve more reliable, high-throughput mycotoxin testing.

Instrumentation Used

• ACQUITY UPLC I-Class PLUS system
• Xevo TQ-S micro triple quadrupole mass spectrometer
• Oasis PRiME HLB pass-through SPE cartridges (3 cc, 150 mg)

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