A simple, robust, sensitive isocratic liquid chromatographic (LC) method suitable for the analysis of spiramycin is described. This method utilizes XTerra RP18 as the stationary phase and acetonitrile-0.2 M K2HPO4 (pH 6.5)-water, 39.5:5:55.5 (v/v), as mobile phase. The mobile phase flow rate is 1.0 mL min(-1); the column is maintained at 70 degreesC by immersion in a water-bath; and UV detection is performed at 232 nm. The method has good selectivity towards the major components (spiramycins I, II, and III), other related substances, and many impurities. The robustness of the method was evaluated by means of full-factorial experimental design.