A human framework needs a careful monitoring of drug concentration which imparts unfavorable condition to human health. Among the various antibiotics, sulfamethoxazole (SMX) has been determined to be a typical antibiotic that is used to treat urinary tract infections, toxoplasmosis, pneumonia, and skin and soft tissue infections. Herein, we report an electrochemical sensing platform using graphene oxide (GO), and nanostructured zinc oxide (ZnO) are used for the detection of SMX. In this study, the working electrodes, GO is modified on FTO using a dip coating technique and on ZnO nanostructures, using a hydrothermal coating method. Finally, GO was dip-coated on FTO in order to obtain a seed layer and the ZnO nanostructure was developed using a hydrothermal method, thus obtaining as GO-ZnO/FTO as a working electrode for SMX detection. Hence, our developed electrodes aimed for high sensitivity that was evidenced to detect SMX at high potential windows. Prior to sensing analysis, the physical and morphological properties of developed electrodes were investigated using X-ray diffraction (XRD) and scanning electron microscopy (SEM) methods. The performance of GO/FTO, ZnO/FTO, and GO-ZnO/FTO was probed by cyclic voltammetry (CV) and current−voltage characteristics (I−V). It was noticed that GO-ZnO/FTO found to exhibit a flower-like structure morphology with effective surface area and active sites for anchoring adsorbate molecules, thus leading to the production of amplified current signals in high potential windows. Under optimized conditions, the designed sensor provided the SMX detection with nanomolar and picomolar limit of detection (0.182 ng/L, 4.8 pg/L, and 0.0949 ng/L), respectively. Besides the specificity, stability and reliability studies were tested for electrodes in multiple real-time water samples for designed sensors. The experimental results of developed sensors show that it has an excellent electron transfer system on the sensor surface among the practical sensors reported for drug applications.