Hazardous mycotoxins such as Ochratoxin A (OTA) can cause negative impacts on both human and animal health and may lead to chronic diseases. Therefore, detection of OTA is important to reduce foodborne disease occurrence. The conventional methods for detection of Ochratoxins has some drawbacks such as time-consuming procedure, high cost and sophisticated equipment. Currently, biosensor with the integration of nanomaterials has becomes a promising tool to overcome the limitations of the previous methods and Graphene is broadly explored due to its high electrical conductivity, expansive surface region and has unique heterogeneous electron transfer rate. Herein, the Water based Graphene paste which is free from harmful organic solvents with good conductivity, and good adhesion with Screen Printed Electrode (SPE) was prepared for Ochratoxin A biosensor. The graphene film is then coated with chemical linkers and biorecognition element. The performance of the stepwise modification of the bioelectrodes was carried out by using electrochemical impedance spectroscopy. Fourier-transform infrared peak shifts clearly shows the successful functionalization of the graphene film. A linear increment trend is observed as the Rct value rises in response to interactions with increasing concentrations of OTA mycotoxin ranging from 1 fg ml-1 to 1 ng ml-1. Overall, this biosensor offers a quick, low-cost, and a simple approach to determine OTA in food and feed samples. © 2023 American Institute of Physics Inc.. All rights reserved.