Identification of impurities in erythromycin by liquid chromatography–mass spectrometric detection

A simple, isocratic liquid chromatographic (LC) method using volatile mobile phase constituents for the identification of related substances in erythromycin samples is described. For method development, evaporative light scattering detection (ELSD) was used. An XTerra™ RP18 column was used at 70 °C with a mobile phase consisting of acetonitrile–isopropanol–0.2 M ammonium acetate pH 7.0–water (165:105:50:680). Mass spectral data were acquired on an ion trap mass spectrometer equipped with an electrospray interface operated in the positive ion mode. First, a library was created using MS/MS and MSn spectra of reference substances available in the laboratory. Using these reference spectra as interpretative templates, eight novel related substances in erythromycin samples were identified: N-demethylerythromycin E, erythromycin E N-oxide, anhydroerythromycin C, N-demethylerythromycin B, anhydro-N-demethylerythromycin A, pseudoerythromycin E enol ether, EF lacking the neutral sugar and EA lacking the neutral sugar.