Inhibition biosensor for determination of nicotine

An amperometric nicotine inhibition biosensor has been substantially simplified and used for determination of nicotine in tobacco sample. Besides the use of single enzyme choline oxidase to replace bienzyme, the use of 1,4-benzoquinone as an electron mediator makes it possible to avoid the use of oxygen or hydrogen peroxide sensor as the internal transducer. Choline oxidase was immobilized on the carbon paste electrode through cross-linking with bovine serum albumin (BSA) by glutaraldehyde. In the presence of choline oxidase and its endogenous cofactor flavin-ademine dinneleotide (FAD), choline was oxidized into betaine while FAD was reduced to FADH2 which subsequently reduced 1,4-benzoquinone into hydroquinone. The later was finally oxidized at a relatively low potential of +450 mV versus saturated calomel electrode (SCE). Nicotine inhibits the activity of enzyme with an effect of decreasing of oxidation current. The experimental conditions were optimized. The electrode has a linear response to choline within 1.25×10^-4 to 1.25×10^-3 mol l^-1. The nicotine measurements were carried out in 0.067 mol l^1phosphate buffer of pH 7.4 at an applied potential of 450 mV versus SCE. The electrode provided a linear response to nicotine over a concentration range of 2.0× 10^-5 to 9.2×10^-4 mol l^-1 with a detection limit of 1.0×10^-5 mol l^-1. The system was applied to the determination of nicotine in tobacco samples.