The purpose of this work was to investigate the effect of l-malic and citric acids and SO 2 on two biochemical properties (diacetyl/acetoin/2,3-butanediol formation and β-glucosidase activity) relevant to flavor development in six Oenococcus oeni strains from wines at pH 4.8 and 3.8. Cells were cultured in MRS without citrate (control medium) and combined with l-malic acid (2 g/L), citric acid (0.7 g/L), and SO 2 (80 mg/L) at pH 4.8 and 3.8. All the test strains grew at all conditions tested including in the presence of SO 2 and at initial pH 3.8, even though growth parameters were maximum in the presence of both the acids at pH 4.8. Organic acids were depleted totally regardless of the condition examined, in which degradation of l-malic acid was faster than that of citric acid. Diacetyl, acetoin, and 2,3-butanediol levels significantly varied depending on the strain for a given condition, for example, at pH 4.8 in control medium the highest value (6.55±0.31 mg/L, strain MS25) represented almost threefold the lowest one (2.43±0.22 mg/L, strain MS9). There was also variability for each strain depending on the initial pH (strains MS25, MS27, and MS48) and the presence of organic acids (all strains except MS25) but not SO 2 . In addition, among strains there was a trend toward mainly diacetyl formation (55%-75%). O. oeni MS9, MS20, and MS46 yielding adequate diacetyl levels were selected for investigating specific β-glucosidase activity and its possible cell localization. Cell suspensions of all the selected strains exhibited positive activities at both pH values which were >4.8. As observed for C4 compounds, organic acids stimulated this activity (28%-49% at pH 4.8; ~20% at pH 3.8), thus partially reverting the inhibition caused by acid stress, while SO 2 did not affect it. The use of different cell fractions (permeabilized cells, cell protoplasts, and cell extracts) associated this activity to the cell surface. Results indicated that diacetyl formation and β-glucosidase activity levels in O. oeni strains as influenced by acidity and organic acids are of relevance for vinification decisions.
Autochthonous Oenococcus oeni strains (MS9, MS20 and MS46) with good malolactic performance and yielding adequate diacetyl levels, were selected to investigate the effect of synthetic and grape glycosides on bacterial growth, substrate utilization and β-glucosidase (βGlu), α-arabinofuranosidase (αAra) and α-rhamnopyranosidase (αRha) activities in a wine-like medium containing 6% ethanol, pH 4.0 (WBM). Then, changes in the volatile compounds profile were evaluated at the end of malolactic fermentation (MLF) carried out by the MS46 strain in WBM containing 1 mg L of natural glycoside. All strains grew and efficiently degraded L-malic acid in WBM where βGlu and αAra activities were found but not αRha. In presence of a synthetic glycoside (eriodictyol 7-O-β-rutinoside) βGlu activity was significantly enhanced for two of the cultures tested (MS20 and MS460) while a low αRha activity was induced, presenting MS46 the better performance. Glycosides extracted from fermented grape musts under different conditions allowed maximum growths, L-malic acid utilization rates and glycosidase activities in the MS46 strain. Thus, βGlu, αAra and αRha activities increased between 30-50 and 3-11% respectively. This indirectly correlated to significant changes in total esters and higher alcohols at the end of MLF, which increased by up to 140 and 30% respectively. Moreover, ethyl and acetate esters formed up to 100-fold than alcohols or esters degraded highlighted the main role of this microorganism in the esters synthesis. Results obtained encourage the potential use of selected indigenous O. oeni strains as a tool to enhance wine complexity through MLF, mainly on highly fruity aroma.
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