Simultaneous screening of multiclass food-borne stimulant-related drug residues in beef using UHPLC-MS/MS

Significance of the Topic

This study addresses the need for robust surveillance of illicit stimulant residues in beef, substances that pose health risks to consumers and violate food safety regulations. Developing a sensitive, reliable, and streamlined analytical workflow enables regulatory laboratories and industry quality control teams to detect and quantify multiple classes of forbidden growth promoters in animal-derived food products.

Aims and Study Overview

The primary goal was to establish a single-method approach for simultaneous determination of 18 stimulant-related drug residues—including β-agonists, anabolic steroids, corticosteroids, and zeranol—in beef muscle. Building on prior work, the authors aimed to simplify sample preparation, minimize lipid-related interferences, and deliver rapid, high-throughput UHPLC-MS/MS analysis suitable for routine screening and confirmation.

Methodology and Instrumentation

Sample Preparation and Cleanup:

• Homogenize 5 g beef, extract with 10 mL acidified acetonitrile (1% acetic acid).
• Salt partitioning using sodium sulfate and sodium chloride, centrifugation.
• Lipid filtration via Captiva nondrip cartridge, vacuum-assisted.
• Desiccation of filtrate with magnesium sulfate, concentration by nitrogen evaporation.
• Reconstitution in methanol/water (7:3) and final centrifugation before injection.

Chromatography and Detection:

• UHPLC system with ZORBAX Phenyl-Hexyl column (2.1×150 mm, 1.8 μm) at 40 °C.
• Mobile phases: 5 mM ammonium acetate/0.01% acetic acid in water (A) and methanol/acetonitrile (7:3, v/v) (B).
• Gradient elution at 0.4 mL/min over 18 min, injection volume 3 μL.
• Triple quadrupole MS with electrospray ionization in dual mode, MRM acquisition of one quantifier and one qualifier transition per analyte.

Main Results and Discussion

Instrument Performance and Separation:
Optimized gradient and column chemistry provided baseline resolution of critical isomer pairs (e.g., betamethasone/dexamethasone) within 18 min.

Linearity and Sensitivity:
All 18 analytes exhibited excellent linearity (R² ≥ 0.995) over 0.10–50.0 μg/L. Limits of quantitation ranged from 0.0020 to 0.30 μg/kg, well below international MRLs.

Matrix Effects and Calibration:
Matrix-matched calibration compensated for minor ion suppression/enhancement. Four compounds showed moderate suppression, one moderate enhancement; external matrix-matched standards ensured accurate quantitation.

Recovery and Precision:
Spike-recovery tests at 0.40, 1.0, and 2.0 μg/kg yielded recoveries of 57–117% (majority within 70–120%), with RSDs of 3–16%, validating method robustness.

Benefits and Practical Applications

• Rapid, high-throughput workflow suitable for regulatory and industrial laboratories.
• Comprehensive multiclass screening in a single analysis.
• Minimal use of organic solvents and reduced labor via lipid filtration cartridge.
• Sufficient sensitivity to meet or exceed current food safety limits.

Future Trends and Potential Uses

The described approach can be extended to other animal tissues (pork, poultry) and additional veterinary drug classes. Ongoing trends include further miniaturization of sample prep, integration with automated SPE–MS platforms, and expansion of target panels to emerging unauthorized growth promoters.

Conclusion

A simple, rapid, and sensitive UHPLC-MS/MS method with lipid cartridge cleanup effectively quantifies four classes of stimulant-related residues in beef. The protocol offers strong linearity, low detection limits, acceptable recoveries, and precision, making it a valuable tool for food safety monitoring.

References

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