The determination of the absolute configuration and enantiomeric composition of chiral species is of great significance. Nevertheless, the accurate quantification through output signals related to the identity of chiral analytes has been rarely reported in this regard. In this study, through the self-assembly of glutathione (GSH) and Cu2+, supramolecular chiral nanoparticles (GSH-Cu2+) were constructed for the enantioselective sensing of chiral 3,4-dihydroxyphenylalanine (DOPA). It is found that GSH-Cu2+ exhibits excellent peroxidase-like activity, with a Kcat value of 2.20 × 10-3 M s-1 g-1 toward 3,3',5,5'-tetramethylbenzidine (TMB). When chiral DOPA is employed as substrate, enantioselective catalysis is observed, with GSH-Cu2+ showing more favorable activity towards D-DOPA as compared to L-DOPA. Further mechanistic study demonstrates that the underlying principle lies in the more favorable adsorption of GSH-Cu2+ toward L-DOPA, which then forms a shielding layer that inhibits the catalytic activity. By exploiting the differential catalytic rates indicated by the ultraviolet-visible (UV-Vis) absorption change of dopachrome (the catalyzed product of DOPA), calibration curves, derived from signals related to their identities, that enable the determination of the absolute configuration and enantiomeric composition of D/L-DOPA, have been established and applied in real pharmaceutical detection. What is also important, the GSH-Cu2+-based sensing platform is demonstrated with high sensitivity (detection limit of 1.31 μM/1.51 μM for D/L-DOPA) and selectivity. Overall, this work enhances our understanding of enantioselective sensing using identity-involved output signals and provides new opportunities for enantioselective sensing with improved reliability.