Analysis of trace residues of tetracyclines in animal tissues and fluids using metal chelate affinity chromatography/HPLC

Abstract

A novel method of analysis for the trace residue determination of tetracyclines in animal tissues and fluids has been developed. Clean-up of sample extracts is based upon the specific ability of tetracyclines to chelate with divalent metal ions (metal chelate affinity chromatography, MCAC) and determination made by high-performance liquid chromatography. The method has been tested for the determination of oxytetracycline (OTC), tetracycline (TC) and chlortetracycline (CTC) in porcine kidney and muscle, ovine kidney, bovine kidney and milk, and trout muscle. Recoveries at the 0.05 mg/kg level for OTC, TC and CTC respectively were 75%, 63%, 73% in porcine kidney, 77%, 79%, 76% in porcine muscle, 85%, 54%, 53% in bovine kidney, 78%, 63%, 57% in ovine kidney, 75%, 58%, 56% in fish (trout) muscle, and 80%, 59%, 59% in bovine milk. At this level both within- and between-batch precision, as measured by the coefficient of variation (CV), was less than 10%. Determination to the 0.01 mg/kg level was carried out in all cases, although the method becomes less precise. The method has been used for several months and found to be both reliable and sufficiently rapid for use as a routine quantitative screening procedure. When coupled with liquid chromatography-mass spectrometry (LC-MS) it is suitable for use as a confirmatory method. Analysis of animals treated with tetracyclines has been carried out.