Journal of Separation Science vol:34 issue:19 pages:2631-2638
A selective reversed-phase (RP) liquid chromatographic (LC) method coupled with UV for the determination of tylosin and its related substances is described. The gradient method uses a Capcell pak C18 ACR column (25 cm x 4.6 mm id, 5 mu m) maintained at a temperature of 60 degrees C. The mobile phases consist of acetonitrile, phosphate buffer pH 5.5 and water: (A; 27.5:10:62.5 v/v/v) and (B; 50:10:40 v/v/v). The flow rate is 1.0 mL/min and UV detection is performed at 280 nm. It allows the separation of all known and 22 other unknown related substances (>= 0.02%) from the main compound and from one another. The method shows good precision, sensitivity, linearity (between 0.2 mu g/mL and 1.25 mg/mL) and robustness. The limit of quantification is 0.2 mu g/mL, corresponding to 0.020%. Seven bulk tylosin samples containing a large number of impurities were examined using this method.