Multiclass residue screening of 217 veterinary drugs in milk and milk powder

Significance of the Topic

Milk and milk powder are staple dietary components globally, prized for their protein, vitamins, calcium, and antioxidants. Veterinary drugs are routinely used in dairy farming to treat infections and enhance productivity. However, improper use can lead to harmful residues in dairy products, posing risks such as allergic reactions, disruption of gut flora, and antibiotic resistance.

Study Aims and Overview

This study presents a comprehensive workflow for the simultaneous screening of 217 veterinary drug residues across 31 chemical classes in liquid milk and reconstituted milk powder. The targeted compounds range from antibiotics (β-lactams, tetracyclines, macrolides, quinolones, sulfonamides) to hormones, NSAIDs, aflatoxins, anthelmintics, and other contaminants. The method aligns with regulatory requirements, offering a high-throughput, sensitive, and reliable solution for quality control in dairy testing.

Methodology

Sample preparation employs protein precipitation extraction with a Na2EDTA-McIlvaine buffer and acetonitrile, followed by lipid removal using EMR-Lipid dSPE and a final cleanup with a polishing kit. Matrix-matched calibration standards at nine levels (0.1–40 ng/g) ensure accurate quantification. Key steps include:

• Reconstitution of milk powder with warm water.
• Extraction of liquid milk using buffer solution and acetonitrile.
• Lipid removal via EMR-Lipid dispersive SPE.
• Acidification and final cleanup with a dedicated polish kit.

Used Instrumentation

The analysis was carried out on an Agilent LC/MS/MS system:

• Agilent 1290 Infinity II LC (binary pump, multisampler, column compartment).
• Agilent Jet Weaver and 20 μL sampling loop.
• Agilent 6495 triple quadrupole mass spectrometer with iFunnel and Jet Stream technology.

Results and Discussion

Method validation in milk demonstrated:

• Linearity: 93% of compounds achieved R2 ≥ 0.990 over 0.1–40 ng/g.
• Recovery: At 5 ng/g spiking level, 86% of analytes fell within 80–120% recovery, with most compounds between 50–150%.
• Repeatability: Over 90% of analytes exhibited %RSD ≤ 15% across three spiking levels. Only two compounds showed %RSD >20% due to low signal intensity.

Representative chromatograms and calibration curves confirmed robust sensitivity and clear separation of analytes in complex milk matrices.

Benefits and Practical Applications

This workflow offers significant advantages:

• High throughput: Simultaneous monitoring of 217 compounds in a single run.
• Broad applicability: Covers diverse chemical classes relevant to food safety.
• Regulatory compliance: Meets international residue limits for dairy products.
• Operational efficiency: Streamlined sample preparation and cleanup reduce analysis time and solvent consumption.

Future Trends and Potential Applications

Emerging opportunities include:

• Expansion to additional matrices such as cheese and yogurt.
• Integration of high-resolution mass spectrometry for non-targeted screening.
• Automated sample preparation systems to further increase throughput.
• Adaptation to monitor new drug classes and emerging contaminants.

Conclusions

The developed LC-MS/MS workflow enables reliable, sensitive, and efficient screening of 217 veterinary drugs in milk and milk powder. Validation results confirm excellent linearity, recovery, and precision, supporting its use in routine food safety monitoring and compliance testing.

References

1. Zeluta A. et al. Antioxidant capacity of cow milk, whey and deproteinized milk. International Dairy Journal. 2009;19:380–385.
2. Han R.W. et al. Simultaneous determination of 38 veterinary antibiotic residues in raw milk by UPLC–MS/MS. Food Chemistry. 2015;181:119–126.
3. Ministry of Agriculture of China. Announcement No. 235: Maximum residue limits in animal-derived food samples. 2002.