Assay of Oxytetracycline in Animal Feeds by Liquid Chromatography with Fluorescence Detection

Abstract

Abstract A liquid chromatographic (LC) method is described to assay oxytetracycline (OTC) in feeds at gram-perton and gram-per-pound levels. Complete feeds and premixes are extracted with acid-methanol, according to AOAC Method 957.23B(i). Extracts are filtered and quantitated by LC using a C18 column and fluorescence detection with excitation at 390 nm and emission at 512 nm. The mobile phase (1.5 mL/min) uses a methanol gradient with an initial hold time of 1 min at 15% methanol followed by an increase to 35% methanol over 8 min. The aqueous component contains CaCI2, disodium ethylenediamine tetraacetate, and sodium acetate buffered to pH 6.5. Recovery of spiked samples averaged 101.6% (coefficients of variation, 0.1- 2.4%), and the limit of detection was 1 g/ton. Results correlated closely with those obtained by using AOAC microbiological Method 968.50 and with another LC method using ultraviolet detection. OTC and epi-OTC elute at approximately 7.4 and 4.8 min, respectively. No interference was observed from several drugs and other antibiotics often used in feeds in combination with OTC