EuroResidue: THE ANALYSES OF MACROLIDES, TETRACYCLINE AND SULFONAMIDE ANTIBIOTICS IN ANIMAL TISSUES USING LC-MS/MS

Importance of the Topic

The monitoring of veterinary antibiotic residues in food products is critical for protecting public health and maintaining consumer confidence. Failure to observe withdrawal periods can lead to residues above maximum residue limits (MRLs), posing risks to human health and complicating international trade.

Objectives and Study Overview

This work presents the validation of two LC-MS/MS methods for quantifying antibiotic residues in animal tissues. One protocol targets macrolide antibiotics in bovine muscle, and a second combines tetracyclines and sulfonamides analysis in shrimp. Both methods were evaluated against EU Commission Decision 2002/657/EC performance criteria.

Methodology

Sample preparation followed standardized procedures for tissue homogenization, extraction, and cleanup to isolate target compounds. Chromatographic separation used reversed-phase gradients at ambient column temperature. Calibration standards were prepared in matrix-matched extracts at concentrations corresponding to MRLs and lower levels to assess sensitivity.

Instrumentation

• Waters ACQUITY UPLC I-Class system with FTN Sample Manager for automated sample handling.
• Waters Xevo TQ-S Micro triple quadrupole mass spectrometer operating in positive electrospray ionization mode.
• Columns:
  • ACQUITY HSS T3, 1.8 µm, 2.1 × 100 mm for macrolide separation.
  • ACQUITY HSS C18, 1.8 µm, 2.1 × 100 mm for tetracyclines and sulfonamides.
• Mobile phases comprised water and organic solvents (acetonitrile or methanol) each containing 0.1% formic acid.

Main Results and Discussion

• Macrolide analysis in bovine tissue achieved baseline separation of 14 compounds within an eight-minute runtime and calibration accuracy (CCa) between 104% and 495% relative to MRLs.
• The shrimp method separated over 20 tetracycline and sulfonamide analytes in under six minutes, with CCa values ranging from 102% to 108% at MRL concentrations.
• Both protocols demonstrated linear response, precision within regulatory limits, and limits of quantification at or below the required MRLs.

Benefits and Practical Applications

• High throughput capabilities allow routine screening of large sample sets in QA/QC laboratories.
• Cost-effective workflows requiring standard UPLC-MS/MS instrumentation ensure broad applicability.
• Validated performance supports regulatory compliance monitoring and due diligence testing by food producers and inspectors.
• Methods can be adapted to additional tissues or species through minor adjustments and revalidation.

Future Trends and Possibilities of Use

Ongoing advancements in mass spectrometry technology, such as higher resolution instruments and enhanced ionization interfaces, are expected to lower detection limits further. Integration with automated sample preparation and data processing software will streamline large-scale residue surveillance. Expanding multi-residue panels to include emerging veterinary compounds and metabolites will enhance comprehensive food safety assessments.

Conclusion

The validated UPLC-MS/MS methods provide rapid, sensitive, and reliable quantification of macrolide, tetracycline, and sulfonamide antibiotics in bovine tissue and shrimp. They meet EU regulatory requirements, support routine monitoring, and offer versatile platforms for extension to other matrices.