Herein, we envisage the fabrication of a highly sensitive enzymatic electrochemical biosensor for selective detection of xanthine (Xn) using xanthine oxidase (XOs) immobilized polyaniline-wrapped titanium dioxide (PANI@TiO2) nanohybrid as a sensing platform. The PANI@TiO2 nanohybrid was synthesized via chemical polymerization using ammonium per sulfate as an oxidant. Various microscopic, spectroscopic, and electrochemical techniques have been utilized to confirm the electrophoretic deposition of the PANI and PANI@TiO2 nanohybrids on to indium tin oxide (ITO) coated glass substrate. The fabricated XOs/PANI@TiO2/ITO electrode exhibits enhanced electron transfer kinetics with an electron transfer rate constant of 0.904cms-1. The electrochemical results show that the fabricated biosensor can detect Xn in the concentration range 1-100µM, with a limit of detection of 0.1µM (S/N = 3) and a response time of 10s. The validation of the biosensors has been conducted using real samples obtained from the rohu (Labeo rohita) fish. The proposed biosensor can be a reliable analytical tool for determining Xn concentration in commercial fish and meat samples.