Chromatographic and Computational Studies on the Chiral Recognition of Sulfated β-Cyclodextrin on Enantiomeric Separation of Milnacipran
Authors: Pathak Pranav , Coutinho Evans C. , Mohanraj Krishnapriya , Martis Elvis , Jain Vikalp 
Published: February 2020

A new, cost-effective and fast chromatographic method using sulfated β-cyclodextrin (SβCD) as a chiral mobile phase additive (CMPA) was developed and validated for the enantiomeric separation of milnacipran. Milnacipran is an anti-depressant drug. Levo-milnacipran is the active enantiomer with less adverse effects than dextro-milnacipran. Hence, it is imperative to separate the enantiomers of milnacipran. Various parameters affecting enantiomeric resolution, for instance, the effect of type and concentration of cyclodextrins, the effect of pH of the mobile phase, effect of type and concentration of the organic solvent in the mobile phase and effect of type of achiral column, were investigated. We demonstrated successful resolution of enantiomers of milnacipran on reverse-phase HPLC with Kinetex C8 column (150x4.6mm, 5µ), using a mobile phase consisting of 18:82 v/v acetonitrile: 10mM sodium dihydrogen orthophosphate dihydrate buffer pH 3.0 (adjusted with orthophosphoric acid) containing 10mM SβCD with a flow rate 1.0 ml/minute. The column temperature was ambient and UV detection was carried out at 227 nm with an injection volume of 20µl. This method for enantiomeric separation of milnacipran was validated in accordance with ICH guidelines and successfully applied to the marketed formulation of Levomilnacipran. Furthermore, molecular docking was used to identify the chiral recognition mechanism. The results of molecular docking corroborated with our experimental findings.A new, cost-effective and fast chromatographic method using sulfated β-cyclodextrin (SβCD) as a chiral mobile phase additive (CMPA) was developed and validated for the enantiomeric separation of milnacipran. Milnacipran is an anti-depressant drug. Levo-milnacipran is the active enantiomer with less adverse effects than dextro-milnacipran. Hence, it is imperative to separate the enantiomers of milnacipran. Various parameters affecting enantiomeric resolution, for instance, the effect of type and concentration of cyclodextrins, the effect of pH of the mobile phase, effect of type and concentration of the organic solvent in the mobile phase and effect of type of achiral column, were investigated. We demonstrated successful resolution of enantiomers of milnacipran on reverse-phase HPLC with Kinetex C8 column (150x4.6mm, 5µ), using a mobile phase consisting of 18:82 v/v acetonitrile: 10mM sodium dihydrogen orthophosphate dihydrate buffer pH 3.0 (adjusted with orthophosphoric acid) containing 10mM SβCD with a flow rate 1.0 ml/minute. The column temperature was ambient and UV detection was carried out at 227 nm with an injection volume of 20µl. This method for enantiomeric separation of milnacipran was validated in accordance with ICH guidelines and successfully applied to the marketed formulation of Levomilnacipran. Furthermore, molecular docking was used to identify the chiral recognition mechanism. The results of molecular docking corroborated with our experimental findings.