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[Display omitted] • A facile and cost-effective polyglycine polymerized pencil graphite electrode (PGl/PGE) was fabricated. • PGl/PGE fabrication was done by simple electropolymerization method. • PGl/PGE was utilized as electrochemical sensor for the determination of Riboflavin (RF). • A low detection limit of 1.24 nM RF was achieved using PGl/PGE. • PGl/PGE may be an efficient in facilitating electron transfer between the electrode and probe molecule. The electronic conductivity and electrocatalytic property of an electrode can be improved by coating its surface with a thin film, polymer or some nanomaterial; however, the fabrication procedure is time-consuming, expensive, and require multistep. In this study, an electrochemically polymerized polyglycine-coated pencil graphite electrode (PGl/PGE) was prepared for the sensitive detection of riboflavin (RF). The formation of glycine polymer on the PGE surface was confirmed by Fourier transform infrared (FTIR), scanning electron microscope (SEM), energy dispersive X-ray spectroscopy (EDX), and elemental mapping analysis. The results of cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) revealed that the charge transfer on PGl/PGE was significantly enhanced when compared to PGE. The fabricated PGl/PGE sensor was used to detect riboflavin (RF) in 0.2 M phosphate buffer (pH 7.0) using square wave voltammetry. The linear concentration range of RF was 2–450 nM under optimized conditions, with a calculated limit of detection of 1.24 nM. Finally, some critical sensor parameters such as sensitivity, selectivity, stability, and reproducibility were evaluated and practically applied for RF detection in pharmaceutical and food samples. [ABSTRACT FROM AUTHOR]