Preparative chromatography was used to obtain approximately 200 mg of each of the four individual isomers of the three methoxytetrahydronaphthalenic derivatives, new agonist and antagonist ligands for melatonin receptors to be tested for binding. The mobile and stationary phases were chosen to achieve best resolution in shorter runtime. Enantiomeric purity was verified and quantified using normal and reversed phase methodology on cellulose CSPs. Enantiomer elution order was analysed and discussed. Limit of detection (LOD) and limit of quantification (LOQ) were calculated.