A solid-state sensor was fabricated by a spontaneous electrochemical deposition of polytyramine (Ptyr) film onto a glassy carbon electrode (GCE) which was further peripherally supported by gold nanoparticles (AuNPs). The surface materials of the developed sensor (AuNPs.Ptyr-GCE) were characterized by X-ray photoelectron spectroscopy (XPS), electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV) and differential pulse voltammetry (DPV). The rate constant of charge transfers (kct) of the as-prepared sensor (8.77 × 10-4 cm/s) was evaluated by fitting the charge transfer resistance (Rct) data in the presence of ferric-ferrous hexacyanide redox couple solution, [Fe(CN)6]3-/4-. The voltammetric behavior of norepinephrine (NOR) was confirmed to follow an irreversible reaction mechanism at which the estimated diffusion coefficient value was 7.39 × 10-5 cm2/s. The sensor showed a large enhancement on NOR oxidation and comparatively lowered its detection limit (DL3s) to 0.130 mM (22 ppb). It was also applied for selective determination of NOR in the presence of high concentrations of ascorbic acid (AA) and uric acid (UA). The interference study highlighted the great stability of the proposed sensor by generating a similar sensitivity as in the pure NOR solution. The analytical performance of the proposed system was validated successfully for pharmaceutical and biological samples with tolerable recovery percentages.