Medical diagnostics and detection of food spoilage require estimation of hypoxanthine (HX), xanthine (XN), and uric acid (UA). A selective sensing platform has been proposed for simultaneous detection of all these species. Functionalized multi-walled carbon nanotube (fMWCNT) stabilized nanogold decorated PEDOT:TOS polymeric nanocomposite (Au-PEDOT-fMWCNT) was synthesized through rapid one-step electropolymerization to enhance conductivity and active surface area by several folds. Electrochemical activities of the proposed sensing platform were analyzed by cyclic voltammetry (CV) and differential pulse voltammetry (DPV), electrochemical impedance spectroscopy (EIS). Analyses through SEM, FESEM and TEM were performed to explore the surface morphology and elemental analysis of the polymeric nanohybrid was investigated by XPS, Raman, FTIR, XRD spectroscopy. Electro-catalysis of UA, XN and HX occurred at low oxidation potentials i.e. 0.082, 0.463 and 0.808 V, respectively in the optimized conditions. The uniquely designed simple, interference free Au-PEDOT-fMWCNT/GCE sensor exhibited high selectivity, good reproducibility, reusability (∼180 times) and stability (∼3 month) with excellent sensitivity of 1.73, 14.31 and 3.82 μA μM-1 cm-2 for UA, XN and HX, respectively. The sensor exhibited linear ranges of detection as 0.1-800, 0.05-175 and 0.1-150 μM with detection limits of 199.3, 24.1 and 90.5 nM for quantification of UA, XN and HX respectively. The performance of the proposed sensor was validated by addition of UA, XN and HX in human serum, urine and fish samples by comparing to those using HPLC. The results indicated good applicability of the proposed sensor for simultaneous detection of UA, XN, HX in real biological fluids.