Poly(p‐aminobenzene sulfonic acid) (pABSA) was electrodeposited onto the surface of a glassy carbon electrode (GCE), which was then used for the preconcentration of l‐tryptophan (l‐Trp) due to the electrostatic and π–π interactions between pABSA and l‐Trp. Polypyrrole (PPy) was electrodeposited onto the surface of the l‐Trp enriched pABSA, and then the l‐Trp templates were removed, resulting in molecularly imprinted PPy/pABSA. To avoid the interference from the oxidation peak of PPy on the following electrochemical chiral recognition of Trp isomers, PPy was overoxidized by cyclic voltammetry (CV). The resultant molecularly imprinted overoxidized PPy (OPPy)/pABSA modified GCE exhibits higher affinity toward l‐Trp than d‐tryptophan (d‐Trp); that is, the oxidation peak current of l‐Trp is greatly higher than that of d‐Trp at the molecularly imprinted OPPy/pABSA modified GCE. [ABSTRACT FROM AUTHOR]