A Robust and Sensitive Method For the Analysis of 510 Pesticides Using LC/MS/MS

Significance of the Topic

Pesticide residues in food pose significant health risks and are tightly regulated by agencies such as WHO, FAO, EPA and the EU. Ensuring compliance with maximum residue limits (MRLs) requires highly sensitive, accurate and high-throughput analytical methods. Multiresidue analysis by LC/MS/MS is a key tool for routine monitoring and quality control in agricultural and food industries.

Objectives and Study Overview

The main goals of this work were to develop, optimize and validate a comprehensive LC/MS/MS workflow for simultaneous quantitation of 510 pesticide compounds across diverse food matrices. The method was verified in multiple laboratories to confirm robustness, transferability and regulatory compliance.

Methodology and Instrumentation

• Instrument configuration: Agilent 1290 Infinity II LC coupled to Agilent 6470 Triple Quadrupole MS in dynamic MRM mode.
• Chromatography: Zorbax Eclipse Plus C18 column (2.1 × 150 mm, 1.8 µm) at 40 °C; mobile phases A = 5 mM ammonium formate in water/0.1 % formic acid, B = 5 mM ammonium formate in methanol/0.1 % formic acid; gradient from 5 % B to 100 % B over 20 min; flow 0.4 mL/min; injection volume 2 µL.
• Sample preparation: QuEChERS extraction followed by dispersive SPE cleanup or Captiva EMR-Lipid cleanup, optimized for high-water (tomato, onion), high-starch (wheat), high-oil (olive oil), high-sugar (honey) and challenging matrices (black pepper).
• Calibration: Seven-point matrix-matched calibration (1–100 µg/L) prepared by spiking blank extracts to account for matrix effects.

Main Results and Discussion

• MRM method development: Automated optimization yielded sharp, well-resolved peaks for 510 targets within the retention time window, demonstrated by a representative chromatogram in honey extract.
• Matrix effect evaluation: 80–120 % response in solvent vs matrix considered negligible; significant effects observed in certain matrices (e.g. >70 % suppression in onion and black pepper), mitigated by matrix-matched calibration.
• Linearity and sensitivity: >90 % of compounds achieved R2 ≥ 0.99; LOD ≤10 µg/L for >99 % of targets in most matrices; LOQ ≤10 µg/kg for ≥82 % of targets in tomato, onion, wheat, honey and olive oil, meeting EU MRLs; black pepper showed ~70 % targets at LOQ ≤10 µg/kg due to ion suppression.
• Precision and recovery: Intra-laboratory precision (RSDr) and inter-laboratory precision (RSDiR) ≤20 % for >90 % of targets; retention time variation within ±0.1 min; average recoveries of 40–120 % for >82 % of compounds across matrices, except black pepper.
• Robustness: Two-day continuous injection of spiked olive oil extract showed RSD <3.5 % for representative pesticides, confirming stable performance of dynamic MRM acquisition.

Practical Benefits and Applications

This workflow enables reliable, high-throughput screening of over 500 pesticides in diverse food commodities. Its transferability across laboratories and compliance with international guidelines streamline routine analysis in QA/QC, regulatory monitoring and research environments.

Future Trends and Opportunities

Emerging directions include expanding compound coverage beyond 750 pesticides via customizable MRM databases, improving extraction efficiency for complex matrices like pepper, integration with high-resolution mass spectrometry for non-target screening, automation of sample prep and AI-driven data review to further enhance throughput and data quality.

Conclusion

The developed LC/MS/MS method provides a robust, sensitive and reproducible platform for quantifying 510 pesticide residues in various food matrices. Validation across multiple labs and compliance with SANTE guidelines demonstrate its suitability for regulatory and industrial applications.

Reference

1. European Commission. Analytical Quality Control and Method Validation Procedures for Pesticide Residues Analysis in Food and Feed, SANTE/12682/2019, 2019.
2. Agilent application note 5994-2370EN, Comprehensive LC/MS/MS Workflow of Pesticide Residues in Food Using the Agilent 6470 Triple Quadrupole LC/MS System, 2020.
3. Agilent application note 5994-3573EN, Analysis of 510 Pesticide Residues in Honey and Onion on an Agilent 6470 Triple Quadrupole LC/MS System, 2021.