Abstract
In this work, a novel voltammetric ethanol biosensor was constructed using alcohol dehydrogenase (ADH). Firstly, alcohol dehydrogenase was immobilized on the surface of a glassy carbon electrode modified by cellulose acetate (CA) bonded to toluidine blue O (TBO). Secondly, the surface was covered by a glutaraldehyde/bovine serum albumin (BSA) cross-linking procedure to provide a new voltammetric sensor for the ethanol determination. In order to fabricate the biosensor, a new electrode matrix containing insoluble Toluidine Blue O (TBO) was obtained from the process, and enzyme/coenzyme was combined on the biosensor surface. The influence of various experimental conditions was examined for the characterization of the optimum analytical performance. The developed biosensor exhibited sensitive and selective determination of ethanol and showed a linear response between 1 × 10−5 M and 4 × 10−4 M ethanol. A detection limit calculated as three times the signal-to-noise ratio was 5.0 × 10−6 M. At the end of the 20th day, the biosensor still retained 50% of its initial activity.
Highlights
Ethanol has been widely used in medicine, biotechnology and the food industry over the years [1].When ethanol concentration reaches toxic levels in fermentation and distillation, it causes inflammation and conjunctiva of the nasal mucous membrane and irritation of the skin
Effect of the cellulose acetate membrane thickness on the electrode response was examined with various ratios of cellulose acetate to covalently attached Toluidine Blue O (TBO)
A novel biosensor based on coimmobilization of TBO, NADH and alcohol dehydrogenase (ADH) on a cellulose acetate coated glassy carbon electrode was developed for ethanol determination
Summary
Ethanol has been widely used in medicine, biotechnology and the food industry over the years [1].When ethanol concentration reaches toxic levels in fermentation and distillation, it causes inflammation and conjunctiva of the nasal mucous membrane and irritation of the skin. Alcohol poisoning occurs at higher ethanol concentration levels. Several analytical methods have been developed so far for the determination of ethanol and other aliphatic alcohols. An alternative way for the sensitive and rapid determination of ethanol is using biosensors based on selective ethanol-converting enzymes [2,7,13,14,15,16,17]. NAD+-dependent alcohol dehydrogenase is selective for primary aliphatic and aromatic alcohols [20]. NAD+ is a very important cofactor since it participates in enzymatic catalysis of more than 300 dehydrogenase enzymes [29,30]. NAD(P)-dependent dehydrogenases are widely used in bioprocesses and analytical applications [3,31,32]. A considerable amount of analytical research has been related to the electrochemistry of the
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