In this study, N and S co-doped chitosan polymer matrix-derived composite (CuS/NSC) was synthesized via a one-step hydrothermal technique using a low-cost copper complex of chitosan polymer. Cyclic voltammetry and chronoamperometry revealed excellent electrocatalytic performance. The glucose sensor exhibited a linear range of 160 μM to 11.76 mM, a low detection limit 2.72 μM and a sensitivity of 13.62 mA mM-1 cm-2 with an excellent linear response. Furthermore, the sensor also displayed selectivity for glucose over potential interfering agents and exhibited a satisfactory recovery percentage using real sample in human serum. The results demonstrate that, CuS/NSC is an efficient nanocomposite material for non-enzymatic glucose sensors and is applicable for glucose detection in biological fluids.