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Poly 1,8-diaminonaphthalene (PDAN) modified electrode based on glassy carbon (GC) electrode was prepared in sulfuric acid (H2SO4) and acetonitrile (ACN) [4.5 M H2SO4 in ACN] mixed solvent using cyclic voltammetry technique (CV). The optimum conditions for the film formation was obtained by sweeping the electrode potential between +0.2 V and +1.2 V with scan rate of 0.1 V s-1 for 20 cycles using 1.0 mM of 1,8-diaminonaphthalene monomer (DAN). Poly 1,8-diaminonaphthalene/GC (PDAN/GC) modified electrode was found to be active in ACN containing 0.1 M lithium perchlorate (LiClO4) and inactive in 0.1 M sodium hydroxide (NaOH). Nickel ions (Ni(II)) were incorporated into the polymer backbone. The obtained new modified electrode with dispersed nickel nanoparticles (Ni/PDAN/GC) exhibits stable redox behavior of Ni(II)/Ni(III) couple in aqueous 0.1 M NaOH solution. Ni/PDAN/GC modified electrode was characterized by CV and scanning electron microscope (SEM). The electro-oxidation of single ascorbic acid (AA), uric acid (UA) and glucose (Glu) at Ni/PDAN/GC modified electrode in 0.1 M NaOH was studied using square wave voltammetry (SWV) technique. Simultaneous determination of AA, UA and Glu at Ni/PDAN/GC electrode in 0.1 M NaOH was achieved where Ni/PDAN/GC electrode showed the capability of potential splitting of oxidation peaks of the present biological compounds. The modified electrode was applied for the determination of UA, AA and Glu in real samples. [ABSTRACT FROM AUTHOR]