Allopurinol (ALO) is a radical scavenging clinical drug, a drug in the treatment of gout, an inhibitor of xanthine oxidase and an effective agent for anti-cancer purposes. The xanthine oxidase is thus essential, and the amount of ALO needs to be controlled more strictly. In this study, a new electrochemical sensor based on magnetite graphene oxide/ordered mesoporous carbon (Fe3O4@GO/OMC) hybrid was prepared and characterized. The results showed sphere shape Fe3O4 nanoparticles with a diameter in the range 17–22 nm on composite. Modification of carbon paste electrode (CPE) with Fe3O4@GO/OMC (Fe3O4@GO/OMC-CPE) allowed the ultrasensitive and selective detection of ALO at oxidation potential of 1.05 V with linear range of 0.05–7 μmol L−1, limit of detection of 47 nmol L−1 and sensitivity of 708 μA mmol−1 L. Also, the results demonstrate that charge transfer at the interface of Fe3O4@GO/OMC hybrid can provide a synergistic effect in comparison with Fe3O4@GO and OMC. The unique surface chemistry of Fe3O4@GO/OMC interface allows π–π stacking and electrostatic interactions with ALO. The advantages are the possibility to regenerate the surface of the sensor, its rapid and easy of production, as well as its applicability for detection of ALO in Tablets and human serum samples, making Fe3O4@GO/OMC-CPE promising interface for bio-electrochemical applications.