In VitroSpecific Activities of Alcohol and Aldehyde Dehydrogenases from Two Flor Yeasts during Controlled Wine Aging

Abstract

Two flor yeast strains of Saccharomyces cerevisiae (S. cerevisiae strains capensis and bayanus) which form velum on the surface of sherry wine during biological aging have been used. Aldehyde and alcohol (isoenzymes I and II) dehydrogenases were detected in vitro during the entire wine-aging process in the flor yeast strains. All enzymatic activities decreased during the first 155 days of wine aging, and after this period, an increase was observed. Ethanol consumption in the wine and the specific activity of alcohol dehydrogenase I were independent of the S. cerevisiae strain. The greater activity of alcohol dehydrogenase II is directly related to the higher acetaldehyde production by S. cerevisiae race bayanus in the wine. This strain has a slower and prolonged growth in the flor film, which permits a continued accumulation of acetaldehyde in the wine. The higher activity of aldehyde dehydrogenase in capensis strain during the flor formation may be related to the production and consumption of large amounts of acetic acid, which involved a more abundant and accelerated cellular growth. We suggest that part of the acetaldehyde might be converted into ethanol during wine aging by alcohol dehydrogenase I in order to maintain a normal redox balance. Keywords: Flor yeast; alcohol dehydrogenase; aldehyde dehydrogenase; biological aging; sherry wine

Effects ofADH2Overexpression inSaccharomyces bayanusduring Alcoholic Fermentation

“…Yeast cells were directly counted under a light microscope in a Thoma chamber. The in vitro enzyme activities of ADH I and II were determined according to the method of Mauricio et al (24). Coenzymes were determined by a high-pressure liquid chromatography-UV method as described by Theobald et al (50) and Mailinger et al (22).…”

“…During biological aging of wine, flor yeasts grow in a medium containing a high ethanol concentration and produce substantial amounts of acetaldehyde (2). The isoenzyme ADH II plays a key role in this process (6,17,24).Overexpression of the ADH2 gene during fermentation can be expected to create conditions of a futile cycle and affect cellular demands for cofactors and also the redox status of the yeast cells.In order to evaluate the impact of overexpression of the gene ADH2 in the V5 wine strain of Saccharomyces bayanus (MATa ura3) during alcoholic fermentation, we cloned the ADH2 gene from S. cerevisiae in a multicopy expression plasmid (pVT100-U) under control of the constitutive promoter ADH1. Overexpression was confirmed by enzymatic analysis and ADH II presence by proteomic analysis of the V5-pVT100-U-ADH2 strain after 48 h of cell growth in a glucose-containing medium and compared to that in the V5-pVT100-U strain used as a control.…”

“…During biological aging of wine, flor yeasts grow in a medium containing a high ethanol concentration and produce substantial amounts of acetaldehyde (2). The isoenzyme ADH II plays a key role in this process (6,17,24).…”

“…Once alcoholic fermentation completes and fermentable sugars are depleted, some S. cerevisiae strains can switch from a fermentative metabolism to an oxidative (respiratory) metabolism and form a biofilm known as "flor" on the wine surface (24). Biological aging is carried out by such yeast strains (flor yeasts) (5).…”

The Microbial Diversity of Sherry Wines

“…Especially, they usually have a considerable number of additional copies of some chromosomes, mainly of chromosome XIII, that contain certain genes related to the oxidative conversion of ethanol to acetaldehyde during biological ageing [24,26]. Flor yeasts are characterized by a high degree of heterozygosis [27].…”