Enantioselective analysis of Moxifloxacin hydrochloride (MOX) enantiomers was investigated on carbon paste electrode (CPE) modified with chiral selector beta-Cyclodextrin and graphene nanosheets (GNS) employing adsorptive stripping differential pulse voltammetric (AdSDPV) technique. Electrochemical chiral discrimination was obtained due to beta-Cyclodextrin, which has 35 chiral centres as a binding site resulting in very tight fit inclusion complex based on host-guest interactions. Enantioselectivity of present electrode system of graphene nanosheets and beta-cyclodextrin modified carbon paste electrode (GNS-beta-CD-CPE) gives difference in anodic oxidation peak potential of enantiomers S, S-MOX and R,R-MOX at 1.039 V and 0.917 V in B.R buffer solution in pH 7 confirming successful enantiomer detection respectively. Various techniques such as X-Ray diffraction (XRD), Scanning electron microscopy (SEM), Thermogravimetric analysis (TGA) and Fourier transform infra red spectroscopy (FT-IR) were employed for characterization of electrode surface. The electrocatalytic response of MOX at GNS-beta-CD-CPE was measured using cyclic voltammetry (CV) and differential pulse voltammetry (DPV). Electrical conductivity of modified electrode was investigated by electrochemical impedance spectroscopy (EIS) which indicates its good conductivity over plain CPE. The proposed enantioselective electrochemical sensor was highly efficient and capable of sensing optically active isomers of MOX which is the S, S-nantiomer from its potential chiral antipode R,R-MOX enantiomer with large difference of 122 mV shift in peak potential which suggests great stereoselectivity is achieved due to chiral environment provided by chiral selector beta-Cyclodextrin. Determination of percentage enantiomeric composition of S, S-MOX in racemic mixture extend future applications of chiral electroanalytical system of beta-CD. In the present work, it is for the first time enantiomeric electrochemical recognition of multi-chiral drug with two chiral centres has been done voltammetrically. (C) 2017 Elsevier Ltd. All rights reserved.