Nucleophilic fraction to estimate the antioxidant activity of inactivated yeast derivates

Abstract

Oxidation in wine is mostly related to the Michael addition of nucleophiles on two quinones formed from the oxidation of ortho-diphenols. In wine this mechanism is responsible for the increase of the yellow hue and aroma loss. Glutathione exerts its antioxidant activity throughout its competitive addition onto quinones, but many other compounds can have the same behavior: sulfanyl compounds, amino acids, etc. Addition of yeast derivates during the winemaking process can increase the level of those nucleophilic compounds and then confer to the wine a higher resistance against chemical oxidation. In order to evaluate the antioxidant efficiency of yeast derivatives the measurement of radical scavenging activity was firstly applied in model wine like conditions. All tested YDs could reduce the DPPH radical, with yeast derivatives enriched in glutathione presenting the highest antiradical capacity compared to those without glutathione enrichment. To estimate the impact of the glutathione concentration on the DPPH results, its concentration was measured on the different solutions. However, there was no clear relationship between the concentration of native glutathione and the anti-radical activity of the YD (spearman correlation ρ = 0.46, p-value > 0.3) despite the known antiradical activity of glutathione. To go beyond the DPPH method, which does not provide any molecular information related to the antioxidant activity of inactivated yeast derivatives, we developed the measurement of inactivated yeast derivatives’ nucleophiles through the evaluation of the specificity and the kinetic of the competitive addition of nucleophilic compounds on the stable quinone 4-methylcatechol. The soluble part of yeast derivatives dissolved in a wine-like a model solution was added to this quinone and thanks to the LC-MS characterization of formed adducts, we could extract this nucleophilic fraction. The pool of 52 nucleophiles other than glutathione enabled to cluster the yeast derivatives according to their initial nucleophilic content and thus their potential antioxidant activity. The DPPH assay revealed the failure of the glutathione concentration to explain the scavenging activity of yeast derivatives soluble fractions. However, the derivatization procedure highlighted the potential of nucleophiles not considered until now to better characterize the antiradical activity of yeast derivatives soluble fractions. This study showed the major importance to consider the global nucleophilic fraction for a better assessment of the antioxidant potential of yeast derivative soluble fractions and highlights the potential of this approach for the characterization of oenological additives.