The most used drug cocktail for the treatment of metastatic or advanced types of cancers is the combination of Doxorubicin (DOX) and Methotrexate (MTX). On one hand, in the chemotherapy, it is crucially important to detect and maintain a good concentration of the anticancer drugs in the patient's body, in order to prevent serious side effects of the drugs. Whereas, on the other hand, these drugs are excreted in faeces and urine from the patient and can contaminate the aquatic environments, which may cause several health hazards. In view of that, an electrochemical sensor was proposed for the simultaneous determination of DOX and MTX using carbon black (CB), cooper nanoparticles (CuNPs), and Nafion modified glassy carbon electrode. Morphological and chemical characterization of these materials was performed using FEG-SEM images and EDS analysis. The modified electrode exhibited good catalytic activity for electrochemical oxidation in square wave voltammetry, with redox potential of 0.69 V and 0.93 V for DOX and MTX, respectively. Under optimized conditions, linear response range of 4.5 x 10(-7) to 5.1 x 10(-6) mol L-1 and 2.2 x 10(-6) to 2.5 x 10(-5) mol L-1 with limit of detection of 2.4 x 10(-8) mol L-1 and 9.0 x 10(-8) mol L-1 were achieved for DOX and MTX, respectively. Finally the CuNPs-CB-Nafion/GCE sensor was successfully applied toward the determination of DOX and MTX in biological matrix (human urine samples) and within the environment (water river samples), where a spike recovers of nearly 100% was obtained.