This paper provides a systematic approach to investigate the reaction of V(IV)/V(V) redox couple and mainly focuses on the effect of surface roughness and functional groups derived from the pre-treatment on the electrochemical behavior of glassy carbon in V(IV)/V(V) solutions. 600-grit, 1200-grit and 4000-grit SiC sandpapers were utilized to change the roughness of the electrode surfaces. Cyclic voltammetry (CV) within different potential ranges were applied in advance to alter oxygen functional groups on the electrode surface as well as surface roughness. CV and electrochemical impedance spectroscopy (EIS) were then used to investigate the electrode behavior in 1 M V(IV)/V(V) (1:1) solution on glassy carbon electrodes. Scanning electron microscopy (SEM) was employed to characterize the surface roughness. X-ray photoelectron spectroscopy (XPS) was adopted to analyze the surface oxygen functional groups. It is shown that polishing and CV scans within different potential ranges can vary the electrochemical response of glassy carbon to V(IV)/V(V) couples significantly. Increased surface roughness is seen to be beneficial for the redox reaction while oxygen functional groups such as C=O and C-O-C tends to reduce the activity of glassy carbon. (C) 2016 The Electrochemical Society. All rights reserved.