QUANTITATIVE DETERMINATION OF VETERINARY DRUG RESIDUES IN EGGS BY UPLC-MS/MS USING A SIMPLE, RAPID AND EFFECTIVE CLEANUP APPROACH

Significance of the Topic

The presence of veterinary drug residues in eggs poses potential health risks, including allergic reactions and antimicrobial resistance. Reliable, multi-residue analytical methods are essential for food safety monitoring and regulatory compliance.

Objectives and Study Overview

This study aimed to develop and validate a simple, rapid cleanup approach for quantitative determination of 17 representative veterinary drugs in eggs by UPLC-MS/MS. Target analytes covered 12 drug classes with established maximum residue limits (MRPLs) in major markets (USA, EU, China).

Methodology and Instrumentation

Sample Preparation and Cleanup:

• Weighed 2.0 g homogenized whole egg.
• Extracted with 8 mL 0.2% formic acid in 80:20 acetonitrile/water (v/v), vortexed 30 s, shaken 30 min, centrifuged at 4 500 rpm for 10 min.
• Passed 1 mL supernatant through Oasis PRiME HLB cartridge (3 cc, 60 mg) at 1–2 psi; no conditioning required.
• Collected eluate and diluted two-fold with 10 mM ammonium formate buffer (pH 4.5).

Chromatography and Mass Spectrometry:

• UPLC system: ACQUITY UPLC I-Class with BEH C18 column (2.1×100 mm, 1.7 µm), column temp 30 °C, flow rate 0.4 mL/min, injection 10 µL.
• Mobile phases: A = 0.1% formic acid in water; B = 0.1% formic acid in methanol; gradient from 15% B to 95% B, total run ~8 min (6.2 min analyte separation).
• MS: Xevo TQ-S triple quadrupole; ESI+, negative for florfenicol; MRM acquisition with optimized transitions.

Results and Discussion

Phospholipid Removal and Recovery:

• The Oasis PRiME HLB pass-through cleanup removed >95% egg phospholipids.
• Recoveries exceeded 80% for 16 of 17 drugs; lasalocid A showed lower recovery.

Linearity, Sensitivity and Precision:

• Matrix-matched calibration curves exhibited excellent linearity (R² ≥ 0.990) over relevant MRPL ranges.
• LODs ranged from 0.5 to 4 ppb, meeting regulatory requirements.
• Accuracy at 0.4×MRL, 1×MRL and 2×MRL levels ranged >70% (except nystatin A1 and lasalocid A); intra-day precision (RSD) was <20% for all analytes.

Benefits and Practical Applications

This streamlined method enables simultaneous quantification of multiple drug classes in eggs with minimal sample handling and efficient phospholipid removal. Its speed and robustness facilitate high-throughput screening in food safety laboratories and routine QA/QC.

Future Trends and Potential Applications

Further developments may include:

• Extension to other food matrices (milk, meat, seafood).
• Automation of sample preparation for increased throughput.
• Integration with high-resolution MS for broader screening.
• Exploration of novel sorbents to enhance recovery for challenging compounds (e.g., lasalocid A).

Conclusion

The proposed UPLC-MS/MS workflow using a one-step Oasis PRiME HLB cleanup offers a simple, rapid, and effective solution for monitoring veterinary drug residues in eggs. The method meets regulatory sensitivity and accuracy requirements for most target analytes and supports reliable food safety assessment.

Reference

• Antonia Garrido Frenich et al. Comparison of several extraction techniques for multiclass analysis of veterinary drugs in eggs using ultra-high pressure liquid chromatography-tandem mass spectrometry. Analytica Chimica Acta, 661 (2010) 150–160.
• The Ministry of Agriculture Bulletin of PRC 235, 2002.
• Global MRL Database. https://www.globalmrl.com/
• Young MS, Van Tran K. Simple and effective cleanup of seafood extracts prior to UPLC-MS/MS multi residue veterinary drugs analysis. Waters Poster, 2015 (PSTR134857156).