Isolation and Characterization of a Trace Level Unknown Impurity of Salmeterol by Chromatographic and Spectroscopic Methods

During the process development of Salmeterol, an unknown impurity was detected at 2.08 Relative Retention Time (RRT) at a level of 0.11% by a gradient Reverse Phase-High Performance Liquid Chromatography (RP-HPLC). The impurity was isolated from the salmeterol drug substance using preparative HPLC. The separation was achieved with an Inertsil C8 column, using acetonitrile–trifluroacetic acid buffer pH 2.7 as mobile phase. The isolated impurity was characterized by NMR and MS techniques. The impurity has been characterized as 4-(2-{[6-(4-cyclohexylbutoxy)hexyl]amino}-1-hydroxyethyl)- 2-(hydroxymethyl)phenol. The synthesis of the impurity and its formation was also discussed