Simplified extraction of tetracycline antibiotics from milk using a centrifugal ultrafiltration device

A simplified method was developed for the simultaneous determination of oxytetracycline, tetracycline, chlortetracycline, and doxycycline in milk. Isolation of the target compounds was performed using an Ultrafree-MC/PL centrifugal ultrafiltration device without prior sample preparation. Analyses were carried out via high-performance liquid chromatography using a Discovery HS F5 column with a gradient mobile phase which consisted of 30 mM citric acid solution (pH 3, in water) and acetonitrile (60:40 → 40:60, v/v). Recovery of the target compounds from spiked samples at four levels (0.05, 0.1, 0.2, and 0.5 μg/mL) was higher than 87%, with relative standard deviations of less than 6%. Limits of quantification ranged from 0.01 to 0.04 μg/mL.