Herein, to fabricate the sponge-like praseodymium oxide nanoparticles (SL- Pr6O11 NPs), a facile and quick sonochemical route is introduced utilizing an environmentally friendly capping agent (Vitis vinifera juice) for the first time. Investigation of the electrochemical behavior of chlorambucil (CHB) showed that the reaction at the surface of the electrode is a diffusion-controlled procedure and the value of the diffusion coefficient (D) was calculated to be 1.35 × 10-5 cm2/s. The suitable electrochemical performance of the Pr6O11 nanoparticles (NPs) enhanced the sensitivity of the sensor for the electrochemical determination of CHB. The sensor signal was linearly proportional to CHB concentration in the range of 8 nM to 220 μM with a detection limit of 1.88 nM under the optimum experimental conditions. Additionally, CHB tablets, human urine and serum samples were successfully used as real samples to detect CHB at SL-Pr6O11/SPE with good recoveries.