Assay and purity control of tetracycline, chlortetracycline and oxytetracycline in animal feeds and premixes by TLC densitometry with fluorescence detection

Abstract

Methods using TLC densitometry with fluorescence detection are described for the assay and purity control of tetracycline (TC), chlortetracycline (CTC), and oxytetracycline (OTC) in animal feeds and premixes. With a silica gel layer previously sprayed with 10% (m/v) sodium EDTA solution adjusted to pH 8.0 or 9.0, all the major impurities were separated from the main components and from each other. The mobile phase consisted of dichloromethane, methanol, and water. After development, the plate was dipped in a 30% (v/v) solution of liquid paraffin in hexane. Quantitation was realized by fluorescence densitometry at 400 nm. The limit of quantitation (LOQ) for tetracycline impurities is 0.8 microg/g, corresponding to 0.2% of the label claimed tetracycline (400 microg/g). The LOQ for impurities of tetracycline and chlortetracycline in premixes is 0.2% of the label-claimed TC (40 mg/g) and CTC (200 or 400 mg/g). The LOQ for impurities of oxytetracycline in a premix is 0.1% of the label claimed OTC (100 mg/g).