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Benznidazole (BZN) is the first‐choice drug for treating Chagas disease (CD). However, it is not ideal for this purpose as it is highly toxic and has irregular pharmacokinetics due to factors such as its low aqueous solubility. These factors necessitate the development of reliable and effective alternative methods for the analytical determination of BZN in biological and pharmaceutical samples. In this context, we present a new electroanalytical method for quantifying BZN using differential pulse voltammetry (DPV) and a glassy carbon electrode (GCE). This method was applied to urine and a pharmaceutical formulation of BZN incorporated into nanostructured lipid carriers (NLC‐BZN). The proposed method provided a linear analytical range of 1.00–10.6 μmol L−1 (R=0.999), with a detection limit of 0.044 μmol L−1 and a quantification limit of 0.13 μmol L−1. The relative standard deviation of the intra‐day and inter‐day precision was below 2.50 %. Through interference studies, the methodology proved to be selective for BZN, because there was no significant potential interference in any of the samples. The recovery tests showed that the accuracy was within the limits recommended in the literature. Therefore, the developed DPV/GCE method can be successfully applied as an alternative method for detecting BZN in NLC‐BZN pharmaceutical formulations and human urine. [ABSTRACT FROM AUTHOR]