Determination of a Single Methodology for the Analysis and Quantitation of Multi- class Veterinary Drugs in Different Animal Matrices Used for Consumption

Importance of the Topic

The monitoring of veterinary drug residues in meat is critical for food safety and regulatory compliance. Multiple classes of pharmacologically active compounds can accumulate in animal tissues and pose health risks to consumers. Regulatory authorities set maximum residue limits (MRLs) at sub-µg/kg levels up to tens of mg/kg or even zero-tolerance for certain substances. A streamlined analytical approach enhances laboratory throughput, reduces costs and ensures consistent quality control in the food industry.

Objectives and Study Overview

This study aimed to develop a single, comprehensive method capable of quantifying over 200 veterinary drugs across different meat matrices (beef, pork, chicken muscle, liver and kidney) within one chromatographic run. In addition to quantitative analysis, the method supports targeted screening using optimized compound databases, facilitating both routine monitoring and investigative testing.

Methodology and Instrumentation

Sample preparation employed a modified QuEChERS protocol: a 2 g homogenized meat sample was extracted with acetonitrile/water (8:2), shaken, centrifuged and filtered through a 0.45 µm PVDF membrane. A 3 µL extract injection was separated on a reverse-phase C18 column under a standardized water/methanol gradient containing 0.1% formic acid. Detection used a triple quadrupole LC-MS/MS system operating in heated electrospray ionization (HESI) for both positive and negative modes. Quantitation utilized selected reaction monitoring (SRM) transitions (3–5 per compound) and optimized collision energies, all managed by dedicated processing software.

Used Instrumentation

• Thermo Scientific UltiMate 3000 RSLC system
• Accucore aQ C18 Polar Endcapped column (100 × 2.1 mm, 2.6 µm)
• Thermo Scientific TSQ Endura triple quadrupole mass spectrometer with HESI source
• TraceFinder software with built-in veterinary drug database (283 analytes)

Main Results and Discussion

Calibration curves in all matrices showed r² values > 0.990 for over 95% of compounds across 12 concentration levels (0.05–300 µg/kg). Limits of detection (LOD) ranged from 0.05–100 µg/kg and limits of quantitation (LOQ) from 0.1–200 µg/kg, meeting or exceeding regulatory MRLs. Some highly polar or poorly ionizing drugs were excluded due to inadequate retention or response. Endogenous compounds such as nicotinamide (100–500 ppb) and riboflavin (> 100 ppb) were detected in all blank matrices, demonstrating the method’s sensitivity and selectivity.

Benefits and Practical Applications

• Consolidation of multiple drug classes in a single analysis reduces sample preparation and instrument run time.
• Robust performance across diverse meat tissues supports routine QA/QC workflows.
• Built-in compound database simplifies method setup and data processing, enhancing reproducibility.

Future Trends and Opportunities

Future developments may include expanding the compound library to cover emerging veterinary drugs and metabolites. Integration with high-resolution mass spectrometry could improve non-targeted screening capabilities. Automation of sample preparation and cloud-based data management will further accelerate high-throughput laboratories.

Conclusion

A multi-class LC-MS/MS method was successfully validated for quantifying over 200 veterinary drugs in various meat matrices. The approach delivers high sensitivity, broad coverage and streamlined workflows, supporting regulatory and quality assurance demands in food safety testing.

Reference

1. EU Regulation 1831/2003/EC; Directive 2009/8/EC
2. U.S. Department of Agriculture, 600 East Mermaid Lane, Wyndmoor, PA 19038