Multiresidue LC-MS/MS Screening of Veterinary Drugs in Milk Using the Ostro Pass-through Sample Preparation Plate

Significance of the Topic

The detection of veterinary drug residues in milk is critical for food safety and regulatory compliance. Rapid, reliable, and high-throughput methods are essential for routine screening to ensure consumer protection and quality assurance in the dairy industry.

Objectives and Study Overview

The study aimed to evaluate the Ostro™ Pass-through Sample Preparation Plate for multiresidue LC-MS/MS screening of diverse classes of veterinary drugs in milk. Key goals included reducing sample preparation time, improving throughput, and achieving effective removal of protein and phospholipid interferences while maintaining acceptable analytical performance.

Methodology and Instrumentation

A streamlined sample preparation protocol was applied to 125 µL aliquots of milk: addition of 0.2 % formic acid in acetonitrile for protein precipitation, filtration and phospholipid retention on the Ostro plate, followed by elution, evaporation, and reconstitution in an aqueous/acetonitrile buffer. Chromatographic separation used a Waters ACQUITY UPLC system with a CORTECS UPLC CSH C₁₈ 1.6 µm, 100 × 2.1 mm column and a gradient of 0.1 % formic acid in water and acetonitrile. Detection was performed on a Xevo® TQ MS operating in positive electrospray ionization with multiple reaction monitoring transitions for tetracyclines, fluoroquinolones, sulfonamides, macrolides, β-lactams, and β-adrenergics. Instrument parameters included a source temperature of 150 °C, desolvation at 500 °C, and a desolvation gas flow of 1000 L/hr.

Results and Discussion

Recovery data at 10 µg/L and 100 µg/L spikes demonstrated satisfactory performance (70–94 % recovery, RSD ≤18 %) for most analytes. Certain tetracyclines and lincomycin fell below quantifiable limits at the lower spike level. Sample preparation throughput was markedly improved: ten samples can be processed in minutes rather than hours. Phospholipid monitoring using the diagnostic 184>184 m/z transition confirmed >98 % removal, minimizing matrix effects and preserving column performance.

Benefits and Practical Applications

• Rapid, walk-away sample preparation integrating protein precipitation, filtration, and phospholipid cleanup.
• High throughput compatible with routine QA/QC and regulatory monitoring.
• Robust LC-MS/MS analysis with sensitive MRM detection across multiple veterinary drug classes.
• Reduced matrix effects and extended column lifetime due to efficient phospholipid removal.

Future Trends and Opportunities

The Ostro plate approach may be extended to other complex food matrices and environmental samples. Integration with high-resolution mass spectrometry could broaden compound coverage and facilitate non-targeted screening. Automation and workflow miniaturization will further enhance throughput and reproducibility, supporting large-scale monitoring programs.

Conclusion

The Ostro™ Pass-through Sample Preparation Plate provides a fast, efficient, and reliable protocol for multiresidue LC-MS/MS screening of veterinary drugs in milk. It enables significant time savings, effective cleanup of proteins and phospholipids, and robust analytical performance suitable for routine laboratory use.

References

1. Young MS, Van Tran K. Optimized Extraction and Cleanup Protocols for LC-MS/MS Multi-Residue Determination of Veterinary Drugs in Edible Muscle Tissues. Waters Application Note 720004144en.
2. HPLC/UV Determination of Tetracyclines in Milk Using Mixed-Mode SPE and eXtended Performance [XP] 2.5 µm Columns. Waters Application Note 720004582en.
3. Contarini G, Povolo M. Phospholipids in Milk Fat: Composition, Biological and Technological Significance, and Analytical Strategies. Int J Mol Sci. 2013;14(2):2808–2831.
4. Eliminating Phospholipids in Drug Discovery Extractions Using a Fast, Generic Sample Clean-up Method. Waters Application Note 720004046en.