In this study, we analyzed the application of potentiodynamic electrochemical impedance spectroscopy (PDEIS) for a selective in situ recognition of biological trace elements, i.e., Cr (III), Cu (II), and Fe (III). The electrochemical sensor was developed using the electropolymerization of aniline (Ani) on the surface of the homemade pencil graphite electrodes (PGE) using cyclic voltammetry (CV). The film was overoxidized to diminish the background current. A wide range of potential (V = −0.2 V to 1.0 V) was investigated to study the impedimetric and capacitive behaviour of the PAni/modified PGE. The impedance behaviors of the films were recorded at optimum potentials through electrochemical impedance spectroscopy (EIS) and scrutinized by means of an appropriate equivalent circuit at different voltages and at their corresponding oxidative potentials. The values of the equivalent circuit were used to identify features (charge transfer-resistant and double layer capacitance) that can selectivity distinguish different trace elements with the concentration of 10 μM. The PDEIS spectra represented the highest electron transfer for Cu (II) and Cr (III) in a broad potential range between +0.1 and +0.4 V while the potential V = +0.2 V showed the lowest charge transfer resistance for Fe (III). The results of this paper showed the capability of PDEIS as a complementary tool for conventional CV and EIS measurement for metallic ion sensing.