Volatile Metabolism of Wine Grape Trincadeira: Impact of Infection with Botrytis cinerea

The aroma of grapes is cultivar dependent and is influenced by terroir, vineyard practices, and abiotic and biotic stresses. Trincadeira is a non-aromatic variety associated with low phenolic content and high sugar and organic acid levels. This cultivar, widely used in Portuguese wines, presents high susceptibility to Botrytis cinerea. This work aimed to characterise the volatile profile of Trincadeira grapes and how it changes under infection with B. cinerea. Thirty-six volatile organic compounds were identified, from different functional groups, namely alcohols, ester acetates, fatty acid esters, fatty acids, aldehydes, and products of the lipoxygenase pathway. Both free and glycosidic volatile organic compounds were analysed by Gas Chromatography and Gas Chromatography coupled to Mass Spectrometry for component quantification and identification, respectively. A multivariance analysis showed a clear discrimination between healthy and infected grapes with 2-trans-hexenal and isoamyl-acetate among the compounds identified as negative and positive markers of infection, respectively. Ester acetates such as 2-phenylethyl acetate, isoamyl acetate, and 2-methylbutyl acetate were present in higher contents in infected samples, whereas the contents of several fatty acid esters, such as ethyl decanoate and ethyl dodecanoate, decreased. These data were integrated with quantitative PCR data regarding genes involved in volatile metabolism and showed up-regulation of a gene coding for Hydroperoxide Lyase 2 in infected grapes. Altogether, these changes in volatile metabolism indicate an impact on the grape quality and may be related to defence against B. cinerea. The presence/absence of specific compounds might be used as infection biomarkers in the assessment of Trincadeira grapes’ quality.

Effect of Must Hyperoxygenation on Sensory Expression and Chemical Composition of the Resulting Wines

This paper evaluates the effect of must hyperoxygenation on final wine. Lower concentrations of caftaric acid (0.29 mg·L−1), coutaric acid (1.37 mg·L−1) and Catechin (0.86 mg·L−1) were observed in hyperoxygenated must in contrast to control must (caftaric acid 32.78 mg·L−1, coutaric acid 5.01 mg·L−1 and Catechin 4.45 mg·L−1). In the final wine, hydroxybenzoic acids were found in higher concentrations in the control variant (gallic acid 2.58 mg·L−1, protocatechuic acid 1.02 mg·L−1, vanillic acid 2.05 mg·L−1, syringic acid 2.10 mg·L−1) than in the hyperoxygenated variant (2.01 mg·L−1, 0.86 mg·L−1, 0.98 mg·L−1 and 1.50 mg·L−1 respectively). Higher concentrations of total flavanols (2 mg·L−1 in hyperoxygenated must and 21 mg·L−1 in control must; 7.5 mg·L−1 in hyperoxygenated wine and 19.8 mg·L−1 in control wine) and polyphenols (97 mg·L−1 in hyperoxygenated must and 249 mg·L−1 in control must; 171 mg·L−1 in hyperoxygenated wine and 240 mg·L−1 in control wine) were found in both the must and the control wine. A total of 24 volatiles were determined using gas chromatography mass spectrometry. Statistical differences were achieved for isobutyl alcohol (26.33 mg·L−1 in control wine and 32.84 mg·L−1 in hyperoxygenated wine), or 1-propanol (7.28 mg·L−1 in control wine and 8.51 mg·L−1 in hyperoxygenated wine), while esters such as isoamyl acetate (1534.41 µg·L−1 in control wine and 698.67 µg·L−1 in hyperoxygenated wine), 1-hexyl acetate (136.32 µg·L−1 in control wine and 71.67 µg·L−1 in hyperoxygenated wine) and isobutyl acetate (73.88 µg·L−1 in control wine and 37.27 µg·L−1 in hyperoxygenated wine) had a statistically lower concentration.

Comparison between Membrane and Thermal Dealcoholization Methods: Their Impact on the Chemical Parameters, Volatile Composition, and Sensory Characteristics of Wines

Over the last few years, the dealcoholization of wine has piqued the interest of winemakers and researchers. Physical dealcoholization methods are increasingly used in the dealcoholization of wines because they can partially or completely reduce the alcohol content of wines. This study aimed to compare the chemical parameters, volatile composition and sensory quality of white, rosé and red wines dealcoholized by two physical dealcoholization methods: reverse osmosis (RO) and vacuum distillation (VD) at 0.7% v/v ethanol. RO and VD effectively reduced the ethanol concentration in all wines to the required 0.7% v/v, but also significantly affected most chemical parameters. The pH, free sulfur dioxide, total sulfur dioxide, and volatile acidity decreased significantly due to dealcoholization by RO and VD, while reducing sugars and total acidity increased significantly. VD resulted in higher color intensity, which was perceptible in dealcoholized rosé and red wines, while RO caused notable color differences in dealcoholized white and red wine fractions. RO were richer in esters (more ethyl esters and isoamyl acetate), higher alcohols, organic acids, terpenics and C13-norisoprenoids, and carbonyl compounds, while wines dealcoholized with VD had lower levels of these volatile compounds, which may reflect both the loss of esters into the distillate during evaporation and condensation (in the case of VD) and a shift in the chemical equilibrium responsible for ester formation and hydrolysis after ethanol removal. β-damascenone exhibited the highest OAV in all wines, however, losses equal to 35.54–61.98% in RO dealcoholized fractions and 93.62% to 97.39% in VD dealcoholized fractions were observed compared to the control wines. The predominant aroma series in the original and dealcoholized wines were fruity and floral but were greatly affected by VD. Sensory evaluation and PCA showed that dealcoholization by RO improved the fruity and floral notes (in rosé and red wines), color intensity, sweetness, viscosity, and aroma intensity better than dealcoholization by VD, while VD mainly enhanced the color of the dealcoholized wines. Both methods increased the acidity of the respective dealcoholized wines. Nevertheless, RO dealcoholized wines achieved higher acceptance by the panelists than VD dealcoholized wines. Therefore, RO may be a better method for producing dealcoholized (0.7% v/v) wines with minimal impact on aroma and sensory quality.

Analyzing the minor volatilome of Torulaspora delbrueckii in an alcoholic fermentation

Torulaspora delbrueckii is an emerging yeast species in the beverage and food industry that is suitable for alcoholic fermentation and to improve the organoleptic quality of wine, beer, mead, and other beverages. Modern consumer preference toward new flavors and products drives the application of T. delbrueckii to ferment less traditional fruits and vegetables. Thus, it has become increasingly relevant to define those metabolites produced in minute quantities by T. delbrueckii, because they may have an impact when producing these new alcoholic beverages. In this study, we have identified metabolites of T. delbrueckii and have compared them with those of Saccharomyces cerevisiae in a controlled setting with a synthetic, high glucose medium using gas chromatography coupled to flame ionization detector (GC–FID) and stir bar sorptive extraction (SBSE) with GC coupled to mass spectrometry (MS). Results showed that T. delbrueckii produced metabolites with higher changes in odor activity complexes than S. cerevisiae: ethyl propanoate, 1,1-diethoxyethane, ethyl isobutyrate, ethyl butyrate, isoamyl acetate, ethyl heptanoate, nonanal, and decanal. We also report seven metabolites detected for the first time in T. delbrueckii. This datum serves to expand the knowledge of T. delbrueckii performance and shows that application of this yeast species is more suitable to a wide array of beverage producers.

Influence of Environmental Microbiota on the Activity and Metabolism of Starter Cultures Used in Coffee Beans Fermentation

Microbial activity is an integral part of agricultural ecosystems and can influence the quality of food commodities. During on-farm processing, coffee growers use a traditional method of fermentation to remove the cherry pulp surrounding the beans. Here, we investigated the influence of the coffee farm microbiome and the resulting fermentation process conducted with selected starter cultures (Pichia fermentans YC5.2 and Pediococcus acidilactici LPBC161). The microbiota of the coffee farm (coffee fruits and leaves, over-ripe fruits, cherries before de-pulping, depulped beans, and water used for de-pulping beans) was dominated by Enterobacteriaceae and Saccharomycetales, as determined by llumina-based amplicon sequencing. In addition, 299 prokaryotes and 189 eukaryotes were identified. Following the fermentation process, Pichia and the family Lactobacillaceae (which includes P. acidilactici) represented more than 70% of the total microbial community. The positive interaction between the starters resulted in the formation of primary metabolites (such as ethanol and lactic acid) and important aroma-impacting compounds (ethyl acetate, isoamyl acetate, and ethyl isobutyrate). The success competitiveness of the starters towards the wild microbiota indicated that coffee farm microbiota has little influence on starter culture-added coffee fermentation. However, hygiene requirements in the fermentation process should be indicated to prevent the high microbial loads present in coffee farm soil, leaves, fruits collected on the ground, and over-ripe fruits from having access to the fermentation tank and transferring undesirable aromas to coffee beans.

A Parameter Study of Ultrasound Assisted Enzymatic Esterification

This work is focused on the study of esterification parameters for the ultrasound assisted synthesis of isoamyl acetate catalyzed by lipase Lipozyme 435 in a continuous loop reactor. Investigating the influence of different parameters shows that a higher concentration of ester (462 mg/gmixture) can be obtained at temperature 50 °C, flow rate 0.16 mL/min. The best ultrasonication conditions are: continuously applied sonication for a short time (20 minutes), ultrasound power 32 mW, and amplitude 20 %. The enzyme can be successfully reused tree times without loss of enzyme activity. Reaction kinetics for isoamyl acetate ultrasound assisted production showed that satisfactory reaction concentration (close to the equilibrium concentrations) could be reached in short reaction times (2 h). Ultrasound assisted enzymatic esterification is consequently a cleaner and a faster process.

Geosmin suppresses defensive behaviour and elicits unusual neural responses in honey bees.

Geosmin is an odorant produced by bacteria in moist soil. It has been found to be extraordinarily relevant to some insects, but the reasons for this are not yet fully understood. Here we report the first tests of the effect of geosmin on honey bees. A stinging assay showed that the defensive behaviour elicited by the bee's alarm pheromone is strongly suppressed by geosmin. Surprisingly, the suppression is, however, only present at very low geosmin concentrations, and completely disappears at higher concentrations. We investigated the underlying mechanisms of the behavioural change at the level of the olfactory receptor neurons by means of electroantennography and at the level of the antennal lobe output via calcium imaging. Unusual effects were observed at both levels. The responses of the olfactory receptor neurons to mixtures of geosmin and the alarm pheromone component isoamyl acetate (IAA) were lower than to pure IAA, suggesting an interaction of both compounds at the olfactory receptor level. In the antennal lobe, the neuronal representation of geosmin showed a glomerular activation that decreased with increasing concentration, correlating well with the concentration dependence of the behaviour. Computational modelling of odour transduction and odour coding in the antennal lobe suggests that a broader than usual activation of different olfactory receptor types by geosmin in combination with lateral inhibition in the antennal lobe could lead to the observed non-monotonic increasing-decreasing responses to geosmin and thus underlie the specificity of the behavioural response to low geosmin concentrations.

Comparison of the Fermentation Activities and Volatile Flavor Profiles of Chinese Rice Wine Fermented Using an Artificial Starter, a Traditional JIUYAO and a Commercial Starter

In this study, an artificial starter culture was prepared using the core microbial species of JIUYAO to produce Chinese rice wine (CRW). The fermentation activity and flavor profiles of CRW samples fermented with traditional JIUYAO, a commercial starter culture, and our artificial starter culture were compared. The optimal protectant combination for lyophilization of the artificial starter was established as 15.09% skim milk, 4.45% polyethylene glycol, 1.96% sodium glutamate, and 11.81% maltodextrin. A comparative analysis revealed that the ethanol content of the three CRW samples was similar. The total acid content of the CRW sample fermented with the artificial starter (7.10 g/L) was close to that of the sample fermented with JIUYAO (7.35 g/L), but higher than that of the sample fermented with the commercial starter (5.40 g/L). An electronic nose analysis revealed that the olfactory fingerprints of the CRW samples fermented with JIUYAO and the artificial starter resembled each other. For both above mentioned samples, the flavor profiles determined by gas chromatography–mass spectrometry indicated some differences in the variety and content of the aroma compounds, but the key odorants (odor activity values ≥1), such as isoamyl acetate, ethyl acetate, phenyl alcohol, and isoamyl alcohol, were similar.

Rational Selection of Mixed Yeasts Starters for Agave Must Fermentation

Tequila and mezcal are both traditional Mexican liquors that are produced from cooked Agave spp. must fermentation and usually rely on spontaneous or pure Saccharomyces cerevisiae strain inoculation. In order to contribute to the rational selection of yeast starters for tequila and mezcal productions, we tested a collection of 25 yeasts originally isolated from mezcal musts, spanning 10 different yeast species. These strains were first characterized in a semi synthetic medium (labeled as M2, having 90 g/L fructose and 10 g/L glucose of initial hexoses) at 48 h of culture, observing a differential pattern in the consumption of sugars and productivity. Selected Saccharomyces strains left around 10 g/L of fructose and showed higher fermentation performance. However, some non-Saccharomyces strains, specifically from Torulospora (Td), Kluyveromyces (Km), and Zygosaccharomyces (Zb) genera, consumed almost all the sugar (i.e., Km1Y9 with <5 g/L) and had a high productivity of ethanol. In general, all Saccharomyces strains presented a high production of ethyl-butyrate, ethyl-decanoate, and ethyl-hexanoate with peaks of 10, 38, and 3 μg/L, respectively. In addition, some Kluyveromyces and Torulospora strains showed a high production of phenyl ethyl acetate (i.e., Km1D5 with up to 1400 μg/L); isoamyl acetate (i.e., Km1D5 and Td1AN2 with more than 300 μg/L), and hexyl acetate (i.e., Td1AN2 with 0.3 μg/L). Representative strains of the most productive genera (Saccharomyces, Torulospora, and Kluyveromyces) were selected to evaluate their fermentative performance and survival in a mixed culture on a medium based on Agave tequilana must, and their population kinetics was characterized using specific fluorescent in situ hybridization (FISH) probes in a qualitative and semi-quantitative analysis during fermentation. We observed that the mixture ratios of 0.1:1:1 or 1:1:1 (Saccharomyces:Kluyveromyces:Torulospora), maintained good fermentation productivities, with alcohol yields above 0.45 g/g, and allowed a high survival rate of the non-Saccharomyces strains during the fermentation process. Finally, mixed inoculum fermentations on A. tequilana must medium, including different Saccharomyces strains and the finally selected Torulospora and Kluyveromyces strains, showed the best production parameters in terms of ethanol, carbon dioxide, glycerol, and acetic acid values, as well as improved volatile metabolite profiles as compared to the pure cultures. All these data were used to propose a methodology of selection of strains to be used as a pure or mixed starter for tequila and mezcal fermentations, with high primary metabolite productivity and desired aromatic profile.