Electrochemical detection of β-nicotinamide adenine dinucleotide (NADH) has fascinated the scientific community due to its significance in pharmaceuticals, food supplements as well as in various biological reactions and pathways. Herein, a facile and low-cost disposable non-enzymatic electrochemical sensor for the selective and sensitive determination of NADH was developed using porous graphene oxide (PGO) modified screen printed carbon electrode (SPE). PGO has received growing attention owing to its high internal porosity, improved conductivity, large surface area, excellent stability, improved active sites, and hence expected to show excellent electrocatalytic activity than graphene and other derivatives of carbon. Firstly, graphene oxide (GO) sheets were synthesized, after which pores were created on these GO sheets through simple base reduction and acid treatment and the pore formation was confirmed using different characterization techniques. The PGO was directly dropcasted on the surface of SPE to form PGO/SPE and the fabricated electrode demonstrated eminent electrocatalytic activity towards NADH oxidation. Further, the fabricated sensor has shown significant response over a broad-range of concentration from 30 to 1071 μM with a good sensitivity of 21.83 μA mM −1 cm −2 and a low detection limit of 8.6 μM. Moreover, the fabricated PGO/SPE has advantages like cost-effectiveness, robustness, disposability, ease of fabrication, high stability, and excellent reproducibility. • Porous graphene oxide was synthesized via base reduction and acidification. • Fabricated a disposable electrochemical sensor by dropcasting PGO on SPE (PGO/SPE) • PGO/SPE effectively utilized for the electrocatalytic oxidation of NADH. • Compared to GO/SPE and ERGO/SPE, PGO/SPE showed superior analytical performance. • PGO/SPE exhibits good selectivity for NADH with excellent stability and sensitivity.