Introduction to Antibiotic Residue Testing

Antibiotics in veterinary use

As well as in applications for human medicine, Antibiotics are used in agriculture for three reasons: to prevent illness, treat sickness and promoteanimal growth when added to feed, although European Union (EU) legislation has forbidden this last practice since 2006¹.

The administration of Antibiotics is not without apublic health risk as Residues may be retainedat toxicological unacceptable concentrations in animal-derived foodstuffs. Antibiotic Residues may also inhibit bacterial cultures in fermentation processes, such as cheese and yoghurt production, and may alter the sensory properties of food products.

Antibiotics may be classified into different groups according to the chemical structure. The families of antibiotics most determined in food (Fig.1) include tetracyclines, phenicols and (fluoro)quinolones, which are currently detected by RnAssays’ Residue Plex test, and β-lactams and sulfonamides, which will be added to Residue Plex in 2011.

Antibiotic resistance

The extensive use of antibiotics has triggered the development of bacterial resistance, which, in recent years, has become an international concern.

Much attention has been paid to food producing animals as potential source of antibiotic resistant bacteria in humans. As a result, there is increasing pressure on laboratories responsible for ensuring the safety of food for human consumption.

Legislation regarding the control of antibiotic residues in live animals and animal products is given in Council Directive 96/23/EC, and maximum residue limits (MRLs) have been established for different matrices. Fast, sensitive methodologies to determine antibiotic residues in food samples are thus critical in food-safety control laboratories.


Confirmatory methods

Confirmatory methods are mostly based on socalled hyphenated techniques such as liquid chromatography(LC) coupled to mass spectrometry(MS), to quantify analyte concentrations. However, they are time-consuming, expensive, and require complex laboratory equipment and trained personnel. Also, they require tedious sample-preparation procedures based on solid-phaseextraction (SPE) and multi-step clean-up.


Screening methods

The ideal characteristics of a screening methodare a very low rate of false-negative samples,high throughput, ease of use, short analysis time,good selectivity and low cost.

Most of the screening methods for antibiotic residue detection in food are based on microbiological and (immuno-)affinity assays. These methods are fast and can be carried out in situ, however they often miss certain antibiotic families including tetracyclines, chloramphenicols and fluoroquinolones.

One of the main problems involved in the analysis of real samples is the antibiotic-extraction procedure,especially from complex matrices (e.g. food). Most extraction procedures employed in reference methods are time consuming and costly.

A remarkable advantage of the screening methods is that the most of them, including ResiduePlex, are based on rapid and simple sample preparation procedures. Speed and simplicity are important characteristics in order to achieve high-throughput methods.

Fig 2 - Examples of antibiotic chemical structures

Microbiological growth inhibition assays are based on a specific reaction between a susceptible organism (generally bacteria) and the antibiotic present in the sample.

Some advantages of these assays are their reliability, cost effectiveness and simplicity. However, the most important drawbacks of the microbiological tests are their lack of specificity and generally the long incubation times required.