The electrochemical modification, surface analysis, and electrochemical impedance spectroscopy of graphite electrodes modified with polymeric films derived from 4-hydroxyphenylacetic acid (4-HPA) were investigated. The electrooxidation and optimization of the immobilization of adenosine monophosphate (AMP) and guanosine monophosphate (GMP) onto poly(4-HPA) films at different pH values was carried out. Variation of the experimental conditions that influenced the electrode reaction, particularly the pH of the electrolytic solution, showed that the oxidation potentials of the immobilized AMP or GMP onto the modified electrodes decreased with increasing pH of the electrolyte. Higher oxidation current was obtained for AMP in phosphate buffer (pH 7.50) solution and GMP in acetate buffer (pH 4.50) solution. Film surface morphology and roughness in the absence or presence of AMP or GMP have been characterized by atomic force microscopy.