Simultaneous determination of ampicillin and tetracycline in milk by using a stopped-flow/T-format spectrofluorimeter

A fast and simple fluorimetric method is described for the simultaneous determination of ampicillin and tetracycline in milk by combining the stopped-flow mixing technique with a T-format spectrofluorimeter. Ampicillin determination is based on its hydrolysis with penicillinase and reaction with mercuric chloride, while the intramolecular energy transfer from tetracycline to europium ions in the presence of thenoyltrifluoracetone is used for tetracycline determination. The similar excitation wavelengths of both systems and the separation between the corresponding emission wavelengths allow the direct determination of both analytes by measuring simultaneously the initial rate of the ampicillin system and the equilibrium signal of the tetracycline system. The relative standard deviations were lower than 2%. The proposed method was applied to the determination of these antibiotics in different milk samples with a recovery of 95.0-103.3% for ampicillin and 93.3-103.3% for tetracycline.