Electrospun titanium dioxide nanofiber/graphite oxide paste/glassy carbon electrode was used as a high sensitive sensor for voltammetric determination of levodopa (L-DOPA) in aqueous media. Electrospinning method was utilized for fabrication of titanium oxide nanofiber. The synthesized nanofibers were characterized by different methods such as TEM, SEM and XRD. The dependence of oxidation peak current on pH of the solution, amount of modifier, scan rate and concentration of analyte were studied to optimize the experimental conditions for electrochemical determination of L-DOPA. The experimental results suggested that the modified electrode promoted electron transfer reaction for the oxidation of L-DOPA. The modified electrode exhibited effective surface area, more reactive sites and excellent electrocatalytic activity toward the oxidation of L-DOPA. In the concentration range of 3 x 10(-7) to 6 x 10(-5) M of L-DOPA, the anodic peak currents, measured by differential pulse voltammetry (DPV), presented good linear relationship with limit of detection (LOD) of 15.94 nM and good sensitivity of 0.0806 mu A mu M-1. The proposed method was successfully applied to quantify L-DOPA in human cerebrospinal fluid, blood serum and plasma. (C) 2014 Elsevier B.V. All rights reserved.