Copper based-metal organic frameworks modified with gold nanoparticles (AuNPs@Cu-MOF) was fabricated via facile approach. The nanocomposite was used to decorate glassy carbon electrode (GCE) for the electrochemical sensing of D- penicillamine (D-PA). The nanocomposite was characterized using different techniques such as scanning electron microscope (SEM), transmission electron microscope (TEM), energy dispersive X-ray spectroscopy (EDX), powder X-ray spectroscopy (PXRD), thermogravimetry (TGA), nitrogen adsorption–desorption isotherms, and Fourier Transform infrared spectroscopy (FTIR). Formation of a new anodic peak of Cu(II)-D-PA complex at + 0.38 V was used to detect D-PA. Cyclic and square wave voltammetric studies proved that AuNPs elevated the conductivity of Cu-MOF. The sensor exhibited wide linear range (0.4–75 × 10⁻⁷ M) and low detection limit (0.11 × 10⁻⁷ M) with a good sensitivity (0.9 ± 0.01 μA/μM). It was successfully applied for the estimation of D-PA in different real samples with recoveries % and relative standard deviations % (RSDs %) in the range of 95.6–102.5% and 2.4–3.3%, respectively.