Racemic indenolol was resolved into its individual enantiomers by high performance liquid chromatography (HPLC) using a commercially available cellulose tris (3,5-dimethylphenyl carbamate) chiral stationary phase, known as Chiralcel OD. Indenolol contains two positional isomers, giving it a total of four enantiomers. The enantiomeric ratio was validated and peak identification for each enantiomer was established according to their optical rotation sign.