Selective determination of methanol and ethanol using a sensor based on alcohol oxidase immobilized on a cassava biopolymer

Abstract

The analysis of alcohol represents an important parameter in food fermentation industries. The use of a biosensor would be an alternative to existing methods. An enzymatic sensor using alcohol oxidase (AOX) was used as the biorecognition element with an immobilized enzyme sensor in combination with an oxygen electrode, optimized in this study to determine the methanol content in cane brandy. The potentiometric signal obtained by oxygen depletion (oxygen consumed) during alcohol oxidation was carried out very quickly in 4.5 s using cassava biopolymer. AOX catalyzes the oxidation of alcohols using molecular oxygen as the electron acceptor, producing corresponding carbonyl compounds and hydrogen peroxide in the reaction medium. An optimal time sensor response to quantify the methanol content of 1.2 s was calculated from the slope of reaction rates.The analysis of methanol with the immobilized enzyme in cane brandy samples revealed very good agreement with the determination performed through standard HPLC methodology. The activity of alcohol oxidase used as a biorecognition element could be wholly retained in a non-dissolvable film (pore size <15 μm) for up to 30 days of storage at refrigeration temperature (4 °C). The sensor was set at a potential of 0.6 mV. A positive linear relationship was found between consumed oxygen as a function of time (mg O2/L × s) and methanol concentration in the range of 0.3–2.5 mM, with a coefficient of R2 = 0.9984 for the immobilized enzyme sensor. This system showed good specificity (KM = 23.5), which continued for twelve weeks during the time of fermentation of the spirit drink.