In this work, a modified electrode named Au/Au NPs-PPy/L-CYs/ZIF-8 was designed and built and simultaneously doped into electropolymerized polypyrrole (PPy) film using cyclic voltammetry (CV). Scanning Electron Microscopy (SEM), Electrochemical Impedance Spectroscopy (EIS), and CV were used to characterize the composite films. The PPy-(ZIF-8) modified Au electrode was used to determine insulin using Square-Wave Voltammetry (SWV). It was found that the prepared zeolitic imidazolate framework-8 had excellent electrocatalytic activity towards insulin oxidation due to its unique properties. The oxidation peak current of insulin hormone increased with its concentration in the range from 1.0 to 60 nM with the linear regression equation: I_pa = 0.3421C (nM) + 3.2762 (γ = 0.998). The measurement limit was estimated to be 1 nM. While the common coexisting substances showed no interference in the response of the modified electrode to insulin, the modified electrode indicated reproducible behavior and a high level of stability during the experiments. The advantages of using these nanocomposites on the surface of modified electrodes include increased stability, good interaction between the analyte and the modified electrode, conductivity, and excellent performance due to the nanometer size of the composites. As a result, it may be particularly suitable for analytical purposes.