Determination of residues of tetracycline antibiotics in animal tissues by high-performance liquid chromatography.

A method has been developed for the determination of tetracycline (TC), oxytetracycline (OTC), chlortetracycline (CTC) and doxycycline (DC) in animal tissues. Tissues were macerated with a buffer solution and centrifuged. The supernatant solution was purified over a Sep-Pak C18 cartridge which had previously been deactivated by silylation. Tetracyclines were eluted with methanol, the solvent evaporated and the residue dissolved in the HPLC mobile phase. Compounds were separated on a Novapak Phenyl Radial-Pak cartridge with a Resolve CN guard column using gradient elution and UV detection. The method was tested on chicken, porcine and bovine muscle. Recoveries were determined for OTC, TC, CTC and DC added to porcine, bovine and chicken muscle at levels of 0.05 and 0.2 mg kg-1. They varied from about 90% for OTC to about 68% for DC with coefficients of variation of 1.8-7.5%. The limit of determination is 5-10 micrograms kg-1. The method was validated on tissues of laying-hens after intramuscular injection of OTC.