Aflatoxin B1 (AFB1) is the most prevalent and toxic contaminant for human beings and animals among all the aflatoxins (AFs). Additionally, AFB1 is documented as a highly carcinogenic and mutagenic contaminant in the literature. Therefore, it is of vital importance to monitor AFB1 contamination to reduce its health associated risks. In the present work, we have designed a label-free electrochemical impedimetric aptasensor for aflatoxin B1 detection. Herein, we compared the analytical performances of two aptamer sequences (seqA and seqB). The detection is based on specific recognition by the aptamer covalently-bound as compact monolayer on screen printed carbon electrodes (SPCEs) via diazonium coupling reaction. The quantification of AFB1 was achieved by using electrochemical impedance spectroscopy. A dynamic quantification range from 0.125 ng mL−1 to 16 ng mL−1 was obtained with both types of aptamer sequences while the detection limits were 0.12 ng mL−1 and 0.25 ng mL−1 for seqA and seqB respectively. For real sample applications, the developed aptasensors were demonstrated in beer and wine samples, and good recovery levels in the range of 92–102% were recorded for AFB1 detection. © 2016 Elsevier B.V.