This paper describes the electrogenerated chemiluminescence (ECL) processes of Ru(bpy)(3)(2+)/nicotine system at ITO working electrode. An ECL-based method for rapid and sensitive detection of nicotine in phosphate buffer solution at pH 8.0 is established. Strong ECL emission was observed at a positive potential of 1.4V vs. Ag/AgCl. A possible ECL mechanism is proposed for the Ru(bpy)(3)(2+)/nicotine system, the oxidation product of nicotine at the electrode surface reacts with the 3+ state of ruthenium bipyridyl (2+) complex and form ruthenium complex exited state ions and thus releases photons. Effect of pH (medium/electrolyte), working potential, buffer composition, buffer concentration, reactant and coreactant (nicotine) concentration, flow rate and loop size on the ECL spectrum of the Ru(bpy)32+/nicotine were studied. At the optimized experimental conditions, lower detection limit for nicotine was observed as 1.2 nmol L-1 (S/N = 3). Linear relationship between ECL current and concentration of nicotine was observed (up to 100 mu mol L-1) with R-value of 0.997. The relative standard deviation with 5 mu mol L-1 concentration of nicotine for 20 analyses was only 1.4%. A 94% recovery rate was observed in a real sample analysis without any complications/disturbance in measurement. Interferences of humid acid, camphor and SDS were not observed in their presence in the sample solution. The established procedure for nicotine quantification manifests fascinating results and can be suggested for further applications. (C) 2008 Elsevier Ltd. All rights reserved.