Application of reversed-phase liquid chromatography and prepacked C18 cartridges for the analysis of oxytetracycline and related compounds

The reversed-phase (RP) chromatographic separation of oxytetracycline (OTC) 4-epioxytetracycline (4-epiOTC), alpha-apooxytetracycline (alpha-apoOTC), and beta-apooxytetracycline (beta-apoOTC) has been accomplished on an Inertsil C8 column at ambient temperature. Using the simplex method of solvent optimization, a 0.1 M ammonium acetate buffer (pH 3.0)-acetonitrile-tetrahydrofuran (72.5:12.5:15, v/v/v) mobile phase was found to give excellent separation of the compounds. OTC, 4-epiOTC, alpha-apoOTC and beta-apoOTC were resolved in 35 min with calculated detection limits of 40, 20, 50 and 140 ng/ml, respectively. Solid-phase extraction (using RP C18 cartridges) of OTC and OTC degradation compounds from distilled water and porcine muscle was tested at four concentration levels ranging from 200 to 2000 ng/ml (g); overall mean recovery of OTC from distilled water and porcine tissue was greater than 90% and 70%, respectively.