Voltammetric investigation of macrolides by an HPLC-coulometric assay

Voltammograms of macrolides, including anhydroerythromycin A, azithromycin, erythromycin A, erythromycin A enol ether, pseudoerythromycin A enol ether, oleandomycin and tylosin have been investigated using a dual electrode cell in combination with a high-throughput LC method. The half-wave potentials (E1/2) of the seven macrolides investigated ranged from 0.734 to 0.866 V, and the current responses reached the maxima at over 1.0 V. The current response of the downstream electrode displayed a non-linear behavior at high potentials over +0.75 V, probably because of polarization of solvent components, e.g., water. The HPLC-coulometric assay was optimized with the potentials of the upstream and downstream electrodes at +0.65 and +0.85 V, respectively. This method is suitable for detection of 14- and 15-membered macrolides (sensitivity < 0.05 μg ml−1), but not for a 16-membered macrolide, tylosin (sensitivity > 0.1 μg ml−1). The assay shows interferences from biomatrices in rat's blood plasma and serum, and human urine, but they were effectively removed by a cold acetonitrile extraction method.