In the present paper, the use of a gold electrode modified by 2-hydroxy-N'1-[(E)-1-(3-methyl-2-thienyl)methylidene] benzohydrazide self assembled monolayer (TMBH) for the determination of Epinephrine (Epi) in the presence of Uric acid (UA) and Acetaminophen (ACOP) was described. Initially, the formation of the TMBH at the surface of gold electrode investigated by electrochemical methods. At the optimum pH of 6.0, the oxidation of Epi occurs at a potential about 28 mV less positive than that of an unmodified gold electrode. The values of electron transfer coefficients (alpha = 0.55), catalytic rate constant (k = 1.2 x 10(-2) M-1 s(-1)) and diffusion coefficient (D = 3.6 x 10(-6) cm(2) s(-1)) were calculated for Epi, using electrochemical approaches. Based on hydrodynamic differential pulse amprometry, the oxidation of Epi exhibited a dynamic range between 3.0 and 100.0 mu M and a detection limit (3 sigma) of 0.08 mu M. Furthermore, simultaneous determination of Epi and UA or ACOP at the modified electrode was described. The Au-TMBH SAM-modified electrode was shown agreeable recoveries in two types of serum; healthy and disease human blood serum. (C) 2012 The Electrochemical Society. [DOI: 10.1149/2.077302jes] All rights reserved.