Fluorimetric determination of tetracyclines in biological materials

Abstract

A method for the fluorimetric determination of tetracyclines in biological materials based on solvent extraction of mixed tetracycline-calcium trichloroacetate ion pairs from aqueous solutions is described. The extraction of tetracycline (TC) and chlorotetracycline (CTC) is almost quantitative, whereas only very poor extraction occurs with oxytetracycline (OTC). However, saturation of the aqueous phase with sodium chloride results in complete extraction of OTC into the organic phase. This effect enables OTC to be determined in the presence of TC and CTC. Ethyl acetate was found to be the most suitable extractant. The fluorescence of the organic phase is measured after addition of magnesium ions and a base.

The excitation maxima of all three tetracycline derivatives are at about 400 nm and emission maxima are at 500 nm. They differ only very slightly and cannot be used for differentiation between the derivatives. The following detection limits for CTC in biological materials were found: serum, 0.05 µg ml^–1; muscle tissue, 0.125 µg g^–1; kidney, 0.15 µg g^–1; liver, 0.3 µg g^–1; and milk powder with high fat content, 2 µg g^–1. Recoveries were between 30 and 45%.