Tetracyclines and Their 4-Epimers

The tetracyclines are a class of broad-spectrum antibiotics, characterized by their hydronapthacene structure, which contains four fused rings. They are widely used in animal 

In addition, parent-only MRLs have been established for DC in beef, swine, and poultry as 100 iig/kg in muscle, 300 iig/kg in liver, 600 iig/kg in kidney, and 300 iig/kg in skin/fat (swine and poultry only). 

The selection of an appropriate method (marker residues, analytical range) therefore depends in part on the regulatory application, given the different regulatory limits applied in different jurisdictions. Since the epimerization of CTC, OTC, and TC is an equilibrium reaction, the epimers are often present to a greater or lesser extent in analytical standards and can occur throughout the extraction and clean-up procedure. Analytical method development is further complicated by the need to separate and quantify parent and epimer, even when the local legislation does not demand it. 

Currently, analytical methods exist for beef, swine, poultry, sheep, and fish tissues as well as for eggs, honey, and milk using various modes of detection. Quantitative methods have used both and more frequently HPLC with 

TABLE 7.5 A Summary of Current Quantitative and Confirmatory MS Methods for Analysis of Tetracyclines and Their Related Degradative Metabolites 

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Biosensors based on microbial immobilization have also been used for food applications, e.g., the inexpensive and rapid high-throughput bacterial biosensor developed by Virolainen et al. for rapid detection of tetracyclines and their 4-epimer derivatives in poultry meat [188, 189]. 

Cherlet M, ScheUcens M, Croubels S, De Backer P, Quantitative multi-residue analysis of tetracyclines and their 4-epimers in pig tissues by high-performance liquid chromatography combined with positive ion electrospray ionization mass spectrometry, Anal. Chim. Acta 2003 492 199-213. 

On the basis of existing knowledge, novel quantitative approaches for the analysis of tetracyclines should involve extraction under mild acidic conditions, incorporate metalchelating agents, and investigate polymeric SPE sorbents. In addition, they should be able to detect and quantify OTC, TC, CTC, DC, and their associated 4-epimers at concentrations that are fit for their intended purpose. If confirmation is required, LC-MS/MS is recommended for its selectivity. 

The stability of various classes at different pH values makes it less than ideal to extract some antibiotics under the same conditions. Tetracyclines are more stable at pH 3-4 than at pH > 5, and changes in pH can lead to the formation of their epimers and even further degradation. Macrolides and f)-lactams are stable at neutral or slightly basic conditions (i.e., pH > 8 or 8.5). Erythromycin degrades to erythromycin-H2O with loss of one H2O molecule at low pH " and is often measured as erythromycin-H20 at m/z 716 in environmental samples but not in food. The... 

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