Abstract
As laccase (produced by Botrytis cinerea) can significantly alter the properties of wine, winemakers frequently use commercially available colorimetric kits and spectrophotometers to measure the activity of this enzyme in grapes, must and wine. Although the used kits are based on electrochemically active substrates (such as syringaldazine and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), ABTS), the electrochemical determination of laccase activity as an alternative to the colorimetric determination was not thoroughly investigated up to now. Therefore, in the present work, we explored the electrochemical determination of laccase activity. Laccase activity measurements were carried out using either carbon fiber microelectrodes or screen-printed electrodes as working electrodes, either syringaldazine or ABTS as the electrochemically active laccase substrate, and either cyclic voltammetry or constant potential amperometry as the electrochemical method. The best performing approach, which combines ABTS, screen-printed gold electrodes, and constant potential amperometry, allowed identifying laccase positive must sample (i.e., must samples with › 3U/mL laccase) in about 5 min.
Highlights
Laccases (E.C 1.10.3.2) are copper oxidase enzymes produced by fungi, plants, and bacteria that catalyze the oxidation of a variety of phenolic and nonphenolic substrates in the presence of molecular oxygen
With laccase activity measurements in a single grape berry in mind, we investigated if carbon fiber microelectrodes (CFMEs) are suitable for the electrochemical evaluation of laccase activity
We examined the performances of screen-printed electrodes for the assessment of laccase activity by chronoamperometry
Summary
Laccases (E.C 1.10.3.2) are copper oxidase enzymes produced by fungi, plants, and bacteria that catalyze the oxidation of a variety of phenolic and nonphenolic substrates in the presence of molecular oxygen. Laccase is one of the metabolites produced in grapes by Botrytis cinerea, a necrotrophic fungus producing significant economic damages in vineyards worldwide. Wines produced from infected grapes suffer enzymatic oxidative processes, irreversibly affecting their organoleptic properties. Detection of Botrytis cinerea in grapes using DNA- or antibody-based methods [1,2] is direct and accurate, but it is complex, time-consuming, and costly, despite recent progress [3,4]. Alternative indirect methods of detecting Botrytis report on indicators such as laccase, gluconic acid, and glycerol, whose production is triggered in infected grapes [2]. While diagnostic methods based on infrared [2]
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