Cells of the yeast Sacchammyces cerevisiae IGC 4072 grown in medium with acetic acid produced a mediated transport system for acetic acid that behaved as an electroneutral proton symport for the anionic form of the acid. The system could transport propionate and formate but not lactate and pyruvate. Uptake of labelled lactic acid was negligible, no mediated transport system activity for this acid being found. The acetate transporter was also found in cells grown in lactic acid or ethanol media, suggesting that the carrier did not require the presence of an external inducer. When cells were grown in lactic acid medium, uptake of labelled acetic acid, a t pH 5.0, was biphasic and consistent with the presence of two distinct transport modes for the acid. One of these components corresponded to the acetatelproton symport, and the higher affinity system corresponded to a more general monocarboxylate carrier that could also transport lactate, pyruvate and propionate. Both systems were subject to repression by glucose, fructose, sucrose, maltose or galactose. In glucose-repressed cells, the undissociated form of the acids appeared to be the only one that could cross the plasma membrane, a diffusion mechanism being involved in the acid uptake. Under these growth conditions and when the extracellular pH was lower than that of the cytosol, accumulation of the acid could also be observed, it being a function of the ApH.

The incomplete consumption of sugar resulting from stuck wine fermentation is associated with important economic losses. One of the solutions to this serious problem consists of reinoculating the brew with a yeast starter culture that is both alcohol tolerant and a vigorous fructose fermenter. The present work aimed to select yeast strains capable of restarting stuck wine fermentations, and identify key parameters that contribute to the efficiency of the strains. Commercial and non-commercial Saccharomyces wine strains were tested, as well as strains of the fermentative non-Saccharomyces species Zygosaccharomyces bailii and Torulaspora delbrueckii. Although the latter species were shown to be more resistant to a combination of ethanol-and acetic-acid-induced cell death, commercial Saccharomyces cerevisiae strains were the most efficient fructose consumers in medium simulating a stuck fermentation. Stationary-phase S. cerevisiae cells performed better than inocula prepared from exponentially growing cultures, which correlates with the higher resistance to ethanol of non-growing populations. Stationary-phase cells pre-adapted to ethanol did not improve fructose consumption rates; this was in contrast to exponential-phase cells that benefited from prior incubation in ethanol-containing medium. Notably, a correlation was observed between yeast fructose consumption capacity and glucose (or fructose) transport. Our results challenge the current belief that ethanol tolerance, expressed in terms of cell viability, is a reliable criterion for the selection of yeast strains to restart stuck fermentations. Instead, this capacity seems to be based on sugar transport and its resistance to ethanol. In an attempt to further improve cell viability in the presence of high ethanol concentrations, hybrid strains of T. delbrueckii and S. cerevisiae were produced, and they showed high potential as restarter strains. The present work opens perspectives for the application of innovative strategies in the wine-making industry.

Acetic acid at concentrations as may occur during vinification and other alcoholic yeast fermentations induced death of glucose-grown cell populations of Saccharomyces cerevisiae IGC 4072 at temperatures at which thermal death was not detectable. The Arrhenius plots of specific death rates with various concentrations of acetic acid (0-2%, w/v) pH 3.3 were linear and could be decomposed into two distinct families of parallel straight lines, indicating that acetic acid induced two types of death: 1) High enthalpy death (HED) predominated at lower acetic acid concentrations (<0.5%, w/v) and higher temperatures; its enthalpy of activation (AH') approached that of thermal death (12.4 x lo4 cal/mol); 2) Low enthalpy death (LED) predominated at higher acetic acid concentrations and lower temperatures with AH' of 3.9 x lo4 cal/mol. While the AH' values for HED induced by acetic acid were similar with those reported earlier for HED induced by other fermentation endproducts, the values for the entropy coefficients were different: 127-168 entropy units mol-'L for acetic acid as compared with 3.6-5.1 entropy units mol-'L for ethanol, which agreed with experimental results indicating that acetic acid is over 30-times more toxic than ethanol with respect to yeast cell viability at high process temperatures.

In the present work the sensitivity of yeast strains of Kluyveromyces marxianus, Pichia anomala, Candida utilis, Schizosaccharomyces pombe and Saccharomyces cerevisiae, to the fungicides cymoxanil, penconazol, and dichlo¯ua-nid, was evaluated. Dichlo¯uanid induced the most negative eects, whereas penconazol in general was not very toxic. Overall, our results show that the parameters IC 50 for speci®c respiration rates of C. utilis and S. cerevisiae and C D for cell viability of S. cerevisiae can be applied to quantify the toxicity level of the above compounds in yeast. Hence, could be explored as an alternative or at least as a complementary test in toxicity studies and, therefore, its potential for inclusion in a tier testing toxicity test battery merits further research. Ó

In glucose-grown cells of Saccharomyces cerevisiae IGC 4072, acetic acid enters only by simple diffusion of the undissociated acid. In these cells, ethanol and other alkanols enhanced the passive influx of labelled acetic acid. The influx of the acid followed first-order kinetics with a rate constant that increased exponentially with the alcohol concentration, and an exponential enhancement constant for each alkanol was estimated. The intracellular concentration of labelled acetic acid was also enhanced by alkanols, and the effect increased exponentially with alcohol concentration. Acetic acid is transported across the plasma membrane of acetic acid-, lactic acid-, and ethanol-grown cells by acetate-proton symports. We found that in these cells ethanol and butanol inhibited the transport of labelled acetic acid in a noncompetitive way; the maximum transport velocity decreased with alcohol concentration, while the affinity of the system for acetate was not significantly affected by the alcohol. Semilog plots of V max versus alcohol concentration yielded straight lines with negative slopes from which estimates of the inhibition constant for each alkanol could be obtained. The intracellular concentration of labelled acid was significantly reduced in the presence of ethanol or butanol, and the effect increased with the alcohol concentration. We postulate that the absence of an operational carrier for acetate in glucose-grown cells of S. cerevisiae, combined with the relatively high permeability of the plasma membrane for the undissociated acid and the inability of the organism to metabolize acetic acid, could be one of the reasons why this species exhibits low tolerance to acidic environments containing ethanol.