Targeted quantitative screening pesticides in food matrices using high resolution DIA spectral library matching

Importance of the Topic

Food safety laboratories require analytical workflows that combine sensitive quantitation with comprehensive screening of pesticide residues in a single analysis. High-resolution LC-MS/MS with data-independent acquisition (DIA) addresses this need by delivering both accurate multi-residue quantitation and confirmatory library matching, in full compliance with EU SANTE 11312/2021 guidelines.

Objectives and Study Overview

The work aimed to establish and demonstrate a generic high-resolution LC-MS/MS DIA method that:

• Quantifies a routine multi-residue pesticide panel in complex food matrices at or below 10 µg/kg.
• Supports extended qualitative screening of additional fungicides via retrospective library searching.
• Meets EU SANTE criteria for mass accuracy (≤5 ppm or ≤1 mDa), retention time tolerance, and ion ratio confirmation.

Methodology and Instrumentation

• Sample Preparation: Standard QuEChERS extraction of spiked matrices (olive oil) with deuterated internal standards (0.1–100 µg/kg).
• Chromatography: Shimadzu Nexera LC with a Shim-pack Velox Biphenyl column (2.1 × 100 mm, 2.7 µm, 40 °C). Mobile phases of water/methanol each containing 2 mM ammonium formate and 0.004% formic acid; flow rate 0.4 mL/min; 4 µL injection with 40 µL water co-injection.
• Mass Spectrometry: High-resolution QTOF (LCMS-9050) in positive ESI mode; TOF MS full scan m/z 140–925 (100 ms); 31 DIA-MS/MS scans (29 ms each) with collision energy spread 5–55 V; isolation widths of 20 Da for m/z 140–540 and 35 Da for m/z 540–925; external mass calibration; total cycle time 1 s.

Results and Discussion

• Quantitation: Over 250 pesticides were quantified at 10 µg/kg in olive oil, complying with EU mass accuracy and ion ratio requirements.
• Library Matching: A curated MS/MS spectral library of more than 500 compounds enabled high-confidence identification based on retention time, accurate mass, isotope pattern, and fragmentation spectra.
• Extended Screening: Retrospective analysis identified additional fungicides (e.g., boscalid, fluopyram, penconazole, pyrimethanil) in grape and nectarine samples beyond the initial target panel.

Benefits and Practical Applications

• Single-method approach reduces instrument time and simplifies method development by supporting both quantitative and non-targeted screening workflows.
• Full compliance with EU SANTE 11312/2021 guidelines ensures reliable reporting and regulatory acceptance in food safety laboratories.
• Retrospective capability allows rapid incorporation of new analytes without additional sampling or reanalysis.

Future Trends and Potential Applications

• Expansion of spectral libraries to include emerging pesticides, metabolites, and degradation products.
• Integration of automated flagging, machine learning, and AI-assisted data processing to further increase throughput and reduce manual review.
• Application of the DIA workflow to other food matrices, environmental samples, and diverse regulatory frameworks beyond EU guidelines.

Conclusion

The described high-resolution LC-MS/MS DIA method delivers a robust, versatile solution for multi-residue pesticide analysis, seamlessly merging quantitation and extensive screening in a single run. It meets stringent EU regulatory requirements, offers high identification confidence through spectral library matching, and enables scalable, retrospective extension to new targets without re-analysis.