Detecting Antimicrobial Compounds Using New UHPLC-MS/MS Technique


Antimicrobials save the lives of millions of humans and animals every year. These substances are often used in farming to treat diseases in livestock that are raised for food. However, misuse is a challenge, particularly in pig farming. Non-compliance with antibiotic label recommendations and withdrawal periods can lead to antimicrobial residues in animal products. Unwarranted drug administration can pose a health risk to consumers, as antibiotic overuse can lead to allergies, food poisoning, disturbances of the intestinal flora, drug resistance, and environmental pollution.

Curious pigs in Pig Breeding farm in swine business in tidy and clean indoor housing farm, with pig mother feeding piglet | Image Credit: © chayakorn -

Curious pigs in Pig Breeding farm in swine business in tidy and clean indoor housing farm, with pig mother feeding piglet | Image Credit: © chayakorn -

Drug resistance problems has led to governments implementing measures to reduce antibiotic use in animals. According to the World Health Organization (WHO), antimicrobial resistance is one of the top 10 global threats to human health. Further, animal agriculture is believed to account for more than half of all antibiotic consumption.

To protect consumer health from adverse effects of veterinary medicines’ presences in food of animal origin, the monitoring of veterinary medicines in food and its derivatives is carried out in the countries of the European Union as part of the National Residue Control Programme. Further, Maximum Residue Limits (MRLs) have been set for most antibacterial substances that can be used in pigs. Under the Rapid Alert System for Food and Feed, seven notifications of veterinary drugs have been registered in Europe in the last five years. All notifications concerned the presence of antibacterial substance above the MRL in pig meat. However, the overall percentage of non-compliant results in pigs is not high, with a report from the European Food Safety Authority EFSA claiming that non-compliant results represented 0.1% of all samples. This has led to a need for new, non-invasive methods that allow ante-mortem control of antibiotics in pigs.

In thist study, the team of scientists examined pig oral fluid is a promising alternative to tissue analysis. Oral fluid collection involves hanging a cotton rope away from water and feed, preventing potential contamination. This approach, in addition to helping avoid residues in edible tissues, can be implemented in an antibiotic residue control program, subsequently preventing economic losses stemming from meat disposal from drugs being detected in tissues post-withdrawal.

For this study, the scientists created an analytical method to analyze 68 compounds from 12 groups of antibiotics (penicillins, cephalosporins, sulfonamides, macrolides, fluoroquinolones, tetracyclines, aminoglycosides, pleuromutilins, diaminopyrimidines, lincosamides, polypeptides, and sulfones) in pig oral fluid. Antibacterial extraction was performed, and analyses were conducted using ultra-high performance liquid chromatography with triple quadrupole mass spectrometry (UHPLC-MS/MS) detection. The method was later validated as a confirmatory method according to the Commission Implementing Regulation (EU) 2021/808, and the reliability of the method was verified by testing real samples from pig farms.

Oral fluid testing can allow for large groups of animals to be tested in a short time, allowing for a safe and non-invasive form of antibiotic control. Further, this approach enables the detection of illegal or inappropriate use of antibiotics in animal breeding, which is vital for food safety and consumer protection. This developed method is novel because no UHPLC–MS/MS method has been described for analyzing a wide range of antimicrobial substances in oral fluid, the scientists wrote. This method can help detect antibacterial residues in edible pig tissue, while also being applied many other veterinary drugs.


(1) Nowacka-Kozak, E.; Gajda, A.; Gbylik-Sikorska, M. Simultaneous Determination of 68 Antimicrobial Compounds in Pigs Oral Fluid by Ultra-High Performance Liquid Chromatography-Tandem Mass Spectrometry. J. Chromatogr. A 2024, 1729, 465053. DOI: 10.1016/j.chroma.2024.465053

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