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Background and Aims Regulations in some wine‐producing countries allow water addition to grape must. The research presented compared the impact of water addition to must on wine phenolics and colour relative to that of wines at the same alcohol concentration, prepared from fruit harvested earlier. Methods and Results Shiraz grapes were harvested at three stages; earlier harvest at 13.4 and 14.4°Be (H‐1, H‐2) and a later harvest at 15.6°Be (H‐3). Water was then added to H‐3 must to match the sugar concentration of H‐1 and H‐2, respectively. Two modes of water addition were followed: (i) water was directly added to the must; or (ii) juice was substituted for water. Both water addition treatments significantly decreased the concentration of phenolics throughout the winemaking process. At the end of malolactic fermentation, the concentration of phenolics, tannin and non‐bleachable pigments, tannin molecular mass and wine colour density were lower in the water addition treatments compared with that of H‐3, but in all other compositional aspects were higher than that of H‐1 and H‐2. Interestingly, where the extent of alcohol reduction by water addition was equivalent, no difference in any of the measured wine attributes was found between the two modes of water addition. Conclusions Water addition to the H‐3 treatment led to higher wine tannin, colour and non‐bleachable pigment than were found in wines made from grapes harvested at a lower sugar concentration. Significance of the Study The study presents a meaningful comparison of potential outcomes in Shiraz wine phenolics and colour for producers who aim to lower alcohol using the approaches of either water addition or early harvest.

Background and Aims Whereas cofermentation is a traditional practice in some Old‐World regions, its outcomes are largely unknown in New‐World wine regions. Herein we report the chemical and sensory effects of cofermentation of Syrah with selected Rhone white cultivars. Methods and Results A Control wine comprising 100% Syrah (SY), and five SY blends [consisting of addition by mass of 10% pressed solids of Viognier (VG), Marsanne (MR), Roussanne (RS), Picpoul (PC), and Grenache Blanc (GB)] were produced. In addition, juice from each white cultivar was fermented separately and blended with finished SY wine at a rate of 10% after malolactic fermentation (MLF). Cofermentation had no effect on wine composition, but blending post‐MLF increased ethanol concentration. Cofermentation resulted in lower concentration of most anthocyanin pigments, whereas blending post‐MLF had the opposite effect. Tannins were higher in SY‐VG, SY‐RS and SY‐MR blends, whereas polymeric pigments increased in SY‐RS post‐MLF and SY‐PC post‐MLF. This was attributed to enhanced copigmentation due to comparatively higher concentration of anthocyanins and flavonols in post‐MLF blended wines, which resulted in a hyperchromic shift at 520 nm. Addition of VG, either as a coferment or as a blend post‐MLF increased the aromatic complexity of the resulting wines. Conclusions Of the white cultivars studied, VG appeared to provide more aromatics both as a coferment or post‐MLF blending option. Post‐MLF blending gave increased aromatics and colour, while offering greater flexibility and fewer logistical issues than cofermentation. Significance of the Study The use of VG as the preferred coferment of SY, and the practice of post‐MLF blending of SY with a small amount of white wine, were validated through chemical and sensory analysis.

Background and Aims Reliable malolactic fermentation (MLF) is essential for process efficiency and spoilage prevention in wine. This study extends previous research in our laboratory, aimed at the development and selection of new bacterial strains for reliable MLF in wine, focusing on ethanol‐tolerant lactic acid bacteria (LAB) strains. Sensory differences of seven LAB strains were assessed, including two commercial strains, two ethanol‐tolerant strains derived from directed evolution and three isolates from a high ethanol Grenache. Methods and Results In this study, the performance of 30 LAB strains was first assessed in fermented chemically defined grape juice media. Seven of the best performing strains were then tested in small‐scale (5 L) fermentations in Shiraz and Shiraz‐Grenache blend wines. All wines were evaluated with a sensory panel using free choice profiling. Conclusions Despite significantly different MLF kinetics between the strains there were no strain‐specific differences on the final wines. The choice of LAB strain did not adversely change the sensory properties of either wine. Significance of the Study These findings provide reassurance that the efficient LAB strains (G71 and G55) and the modified directed evolution strains do not compromise the sensory properties of wines despite their marked MLF benefits.

Background and Aims The encapsulation of Oenococcus oeni into silica‐alginate (Si‐ALG) gels has been previously confirmed as a suitable strategy for a successful malolactic fermentation (MLF). The aim of this study was to evaluate the effect of two inoculation strategies (simultaneous vs sequential) on the performance of MLF with encapsulated O. oeni under winemaking conditions. Methods and Results Sequential inoculation of Si‐ALG biocapsules successfully achieved a complete MLF in high ethanol wines, while free and ALG encapsulated bacteria failed. A simultaneous inoculation with Si‐ALG biocapsules provided a significant reduction in time to complete MLF in high sugar and low pH musts. The regime of inoculation did not modify the chemical composition of the wines. Conclusions Either sequential or simultaneous inoculation of Si‐ALG biocapsules constitute an effective alternative to free bacteria to undertake MLF under stressful conditions. Significance of the Study Effective winemaking protocols based on sequential or simultaneous induction of MLF with inoculated Si‐ALG encapsulated O. oeni have been accomplished. These are of potential interest for winemaking in both warm and cool regions.

The genus Pediococcus comprises a group of nutritionally fastidious bacteria, several of which have been isolated from wines worldwide, for example P. damnosus, P. inopinatus, P. parvulus and P. pentosaceus. The growth of these bacteria is generally considered undesirable because of their ability to produce off‐odours and flavours. As examples, Pediococcus spp. have been associated with the synthesis of excessive diacetyl, exopolysaccharides, and/or biogenic amines, all of which have a detrimental impact on wine quality. Historically, oenological factors, such as low pH, high ethanol and nutrient depletion following malolactic fermentation, have often been considered sufficient to limit growth. Recent research, however, has supported the contention that Pediococcus spp. can grow in wines considered to be microbiologically stable. Furthermore, the presence of Pediococcus spp. in wines does not always lead to spoilage. New findings have suggested potential uses for Pediococcus spp. in winemaking, but have also highlighted a lack of understanding of those factors that influence growth and spoilage potential. The present work reviews the current knowledge of pediococci isolated during vinification as well as focuses on areas for future research.

Background and Aims Malolactic fermentation (MLF), accomplished by lactic acid bacteria (LAB), is indispensable for the production of some red wines. As the key additive for wines, sulfur dioxide (SO2) frequently affects LAB growth and MLF development. We aimed to investigate suitable reference genes in Lactobacillus plantarum R23, an excellent LAB with good tolerance to SO2, for a real‐time quantitative PCR analysis of SO2 tolerance. Methods and Results The expression of eight candidate reference genes under SO2 stress conditions for L. plantarum R23 was analysed, and geNorm and BestKeeper software packages were employed to assess the reference genes. The expression of mleR1 and mleP1 under SO2 stress conditions was used to confirm the results. The geNorm analysis indicated that the stability ranking of reference genes was rpoB > rpoC > recA > ldh > cfal > mtlR > asd2 > gpp. Both software packages suggested that four genes should be selected for accurate normalisation, and the validated experiment further confirmed the suitability of the reference genes revealed in this study. Conclusions The genes rpoB, rpoC, recA and ldh were the most stably expressed reference genes and thus represent the reference gene combination that should be used to obtain the most accurate results for different SO2 stress conditions in L. plantarum R23. Significance of the Study This is the first detailed study to evaluate selected reference genes in L. plantarum R23 under SO2 stress conditions, which should allow the quantification of bacterial gene expression levels under SO2 stress conditions and also provide a foundation for the more accurate use of real‐time quantitative PCR in L. plantarum.

Background and Aims This study evaluated the impact of pre‐fermentative cold maceration (PCM), in the presence of two non‐Saccharomyces yeasts, Metschnikowia pulcherrima MP 346 and Metschnikowia fructicola MF 98‐3, and of a commercial pectic enzyme, on fermentation kinetics and on the volatile composition of a Sangiovese red wine. Methods and Results Sangiovese grape must was inoculated with MP 346 or MF 98‐3 or treated with a pectic enzyme preparation during PCM, at 5°C for 24 or 72 h. A Control wine was produced by a pure culture of Saccharomyces cerevisiae. Both non‐Saccharomyces strains affected the initial yeast population dynamics and the persistence of S. cerevisiae at the end of malolactic fermentation. Irrespective of the duration of PCM, the inoculum of Metschnikowia strains did not influence the rate of sugar consumption or the kinetics of malolactic fermentation. The volatile composition of the final wines was evaluated with solid‐phase extraction, followed by GC/MS. The concentration of some terpenes and C13‐norisoprenoids, nerol, geraniol, 8‐hydroxy‐linalool (cis) and 3‐oxo‐α‐ionol, and of some esters, isoamyl lactate and ethyl isoamyl succinate, was higher in wines inoculated with Metschnikowia strains than in the Control and wine treated with pectic enzyme. Conclusions Metschnikowia yeast strains MP 346 and MF 98‐3 affect wine volatile composition. Significance of the Study This study shows for the first time that an inoculum of Metschnikowia strains MP 346 and MF 98‐3 during PCM is effective in modulating the volatile composition of a Sangiovese red wine.

Background and Aims The concentration and composition of nitrogen (N) in must is critical to yeast fermentation and aroma profiles in wine. We investigated the effect of adding branched-chain amino acids (BCAAs, including l-valine, l-leucine and l-isoleucine) to a non-limited N medium on the formation of aromatic compounds and on gene expression patterns of the wine yeast Saccharomyces cerevisiae. Methods and Results A synthetic N-sufficient must was supplemented with BCAAs and fermented with S. cerevisiae. The fermentation kinetics, aroma characteristics and yeast global gene expression in wine with addition of BCAAs were compared with that of the Control. Addition of BCAAs improved yeast growth and fermentation activity, increased the production of higher alcohols, medium-chain fatty acids and their corresponding esters (82, 34 and 49% increment compared to that of the Control, respectively). The comparative transcriptomic analysis revealed that 25 genes varied in their expression during fermentation with added BCAAs relative to that of the Control, which are mainly involved in yeast growth and proliferation (GIP1, DIT2, RCK1, SPO1 and MEK1) and transport of amino acids (GAP1). Conclusions An increase in BCAA transport ability and in yeast population is the main reason for achieving an increased concentration of volatile aroma compounds in wine with addition of BCAAs. Significance of the Study Our results suggest that adding BCAAs into N-sufficient must is still an efficient way to adjust wine's aromatic composition and improve aroma complexity.

Background and Aims Beginning and completion of malolactic fermentation (MLF) are complicated because of the harsh environment of wine. Encapsulation of lactic acid bacteria could be an attractive strategy to improve their activity and stability. This work evaluates the fermentative capacity of Oenococcus oeni encapsulated into silica–alginate (Si–Al) and alginate capsules under adverse winemaking conditions (high alcohol concentration, low pH and low temperature). Mechanical and chemical stability of capsules was also monitored. Methods and Results Silica–alginate capsules showed higher malolactic activity and better mechanical and chemical stability than untreated alginate capsules. Their capacity for bacteria retention was similar to that of alginate capsules. Ethanol, pH and fermentation temperature had great influence on fermentative behaviour of free and encapsulated bacteria. Under extreme winemaking conditions, such as high ethanol concentration (13–16%), low pH (3.0–3.3) and low fermentation temperature (13–15°C), the highest consumption of malic acid was achieved using Si–Al capsules. Free and encapsulated bacteria showed similar fermentative profiles in the presence of variable concentration of malic acid and nitrogen in wine. Conclusions Encapsulated O. oeni in Si–Al gels enhanced MLF in wines with a high concentration of ethanol and low pH and at low fermentation temperature. Moreover, inclusion of an inorganic network of silicates significantly improved the mechanical robustness and chemical stability of alginate capsules. Significance of the Study The application of Si–Al-encapsulated bacteria is an interesting strategy to improve MLF in warm climates where wines have higher ethanol concentration and in cool climates with higher acidity and low fermentation temperature.

Background and Aims The aim of this work was to assess and compare the chemical composition and colour characteristics of Barbera red wines obtained after partial alcohol reduction using three promising methodologies for implementation at the industrial level. Methods and Results Alcohol reduction was achieved by: (i) pre-fermentation addition of liquid derived from grape must (reverse osmosis by-product); (ii) mixed fermentations with strains of Starmerella bacillaris and Saccharomyces cerevisiae; and (iii) dealcoholisation of wine post-fermentation with a polypropylene membrane. The microbiological approach enabled the production of wines with a slightly lower alcohol concentration (−0.2 to −0.3% v/v), while facilitating the release of anthocyanin and some esters of fatty acids (ethyl hexanoate and ethyl dodecanoate) that could contribute positively to wine aroma with pleasant nuances. The low impact of the partial replacement of grape juice on the chemical composition and chromatic characteristics of Barbera wines makes this technique a good option for reducing the ethanol concentration by up to 1.0–2.0% v/v. In contrast, the use of a polypropylene membrane influenced negatively the composition of red wines by reducing significantly the concentration of esters (−60%) and anthocyanin (−17%), independently of the dealcoholisation level (up to −2% v/v). Conclusions The alcohol reduction strategies can greatly influence the volatile and phenolic composition of the wine. The choice of either a technological or microbiological approach in the wine industry is dependent on the alcohol reduction required. Significance of the Study This is the first comparative study of three strategies to reduce the alcohol concentration on the same batch of must/wine.

Background and Aims The light-struck taste is a fault occurring in light-exposed white wine containing methionine and a high concentration of riboflavin (RF) and bottled in clear bottles. These conditions induce the formation of methanethiol and dimethyl disulfide, responsible for a cabbage-like aroma. In order to decrease the risk of wine spoilage, a low concentration of RF should be obtained in wine either by preventing RF release from yeast during winemaking or by removing RF from wine. Methods and Results Fifteen commercial Saccharomyces strains intended for the wine industry were tested for RF production, which was also evaluated when two yeast-based nutrients were added into the must for one of these strains. The RF released during vinification was strain-dependent and a concentration from 30 to 170 μg/L was found in wine. A high concentration of RF was released in the presence of the yeast-based nutrients because of either the yeast metabolism or the RF contained in the nutrient itself. The ability of different inorganic (bentonite, charcoal, zeolite, kaolin) and organic adjuvants (egg-white proteins, polyvinylpolypyrrolidone) to deplete RF in wine was evaluated. A relatively low level of charcoal (50 mg/L) removed up to 60% of RF in wine, although its effectiveness was related to the charcoal source. A high concentration of bentonite (1 g/L) was needed to effectively decrease the risk of wine spoilage. Conclusions A critical RF concentration in white wine can be prevented by applying one or more approaches in winemaking: using low RF-producing yeast strains in fermentation, selecting suitable yeast nutrients or adsorbing RF by insoluble charcoal or bentonite. Significance of the Study The research highlights suitable tools to prevent light-struck taste in white winemaking.

Background and Aims This study evaluated the impact that the non-Saccharomyces yeasts, Torulaspora delbrueckii (TD), Lachancea thermotolerans (LT), Pichia kluyveri (PK), Metschnikowia pulcherrima (MP), Candida zemplinina (CZ) and Kazachstania aerobia (KA), in sequential inoculation with Saccharomyces cerevisiae (SC), had on the volatile chemical profile of Shiraz wine. Methods and Results Shiraz grape must was inoculated with monocultures of the non-Saccharomyces yeasts, which were allowed to ferment until 2% v/v ethanol concentration was reached at which point, SC was added to complete the alcoholic fermentation. The control was SC in monoculture. The final wines were subjected to solid phase microextraction-GC × GC-time of flight-MS to evaluate the untargeted volatile metabolite profile of each treatment. Each fermentation produced a unique chemical profile. The LT–SC sequential fermentation was the most significantly different from the control primarily in the ester, alcohol and terpene profile. The KA–SC sequential fermentation had the highest amount of volatile acidity, and the PK–SC sequential fermentation had a relatively high amount of acetaldehyde and a few esters. The MP–SC sequential fermentation also revealed a higher concentration of several esters. The TD–SC sequential fermentation was notable for its lack of a distinct pattern in comparison with that of the other fermentations. Conclusions Given these characteristics, the LT–SC sequential fermentation showed the most potential for increased chemical complexity of the Shiraz volatile profile. Significance of the Study The results demonstrate that there is no clear, singular trend for how different, non-Saccharomyces species of yeast – once thought to be wholly unfavourable in wine fermentations – will impact wine chemistry, flavour or quality. Each species presents unique metabolic characteristics, many of which could be beneficial, and their potential application in the wine industry should be considered.

Background and Aims The request by the consumer for safer food is also pushing the wine sector to find alternative solutions to sulfur dioxide. The aim of this research has been to evaluate the application of ozone as a sanitising agent of grapes before vinification in order to avoid the use of sulfur dioxide. Methods and Results Postharvest ozone fumigation overnight of Petit Verdot grapes increased anthocyanin concentration by 19% versus 9% in the Control (untreated) during fermentation/maceration. Fermentation kinetics were faster in wine made from ozone-treated grapes (16 vs 21 days for the Control), and extraction of phenolic substances and anthocyanin was more rapid than that in the Control wines, Ozone treatment significantly reduced the count of Saccharomyces cerevisiae and non-Saccharomyces yeasts but also significantly decreased the acetic acid bacteria. The final wine produced from ozone-treated grapes was characterised by low volatile acidity (similar to that of the Control wine) with a significant reduction in sulfur dioxide (17 and 8 mg/L, respectively, total and free sulfur dioxide). Sensory evaluation revealed a strong fruity aroma. Conclusions Ozone gas treatment of grapes reduced the microbial count significantly and increased the extraction of phenolic substances and the aroma of the final wine. Significance of the Study Postharvest ozone fumigation can be used to produce wine without sulfur dioxide.

Background and Aims Oenococcus oeni is the lactic acid bacteria species that best adapts to harsh wine conditions. This species is currently the main component of the malolactic starter cultures used in winemaking. Other species of lactic acid bacteria, however, such as Lactobacillus, can also conduct the malolactic fermentation, especially in low acidity wines. This study aimed to identify suitable Lactobacillus strains and inoculation methods to undertake the malolactic fermentation in wines with pH > 3.5. Methods and Results Six Lactobacillus strains of species L. mali (E4634), L. paracasei (E4539, E4541), L. plantarum (E4538, E4608) and L. satsumensis (E4555) were selected for their good growth performance and high malolactic activity in grape must, although these characteristics differed among them. Freeze-dried starter cultures were obtained for strains E4538, E4608, E4555 and E4634. These starter cultures showed a high rate of malic acid consumption in grape must. Inoculation of the Lactobacillus strains in grape must, prior to fermentative yeasts, was selected as the best inoculation strategy to promote the malolactic fermentation. Conclusions Inoculation of the starter cultures of the selected Lactobacillus strains before yeast inoculation in grape must effectively allows malolactic fermentation in wines. Significance of the Study Inoculation of Lactobacillus strains in grape must, prior to fermentative yeasts, is an effective alternative to O. oeni to undertake malolactic fermentation in wines of pH > 3.5.

Background and Aims Commercial enzyme preparations used in winemaking are mixtures of several enzymatic activities, such as pectinases, cellulases and hemicellulases. The aim of this work was to evaluate the side activities of commercial enzyme preparations and their collateral effects on white wine. Methods and Results The β-glucosidase, cinnamoyl esterase and protease activities of 15 enzyme preparations currently available in the market were screened. Must was treated with the enzyme preparations, and the collateral effects on white wine were evaluated. The application of cinnamoyl esterase-rich preparations induced an increase in hydroxycinnamic acids and 4-vinylguaiacol in the wine that reached a final concentration about twice that of the no enzyme control. Cinnamoyl esterase activity, above 50 U/g or mL of preparation on chlorogenic acid, leads to the risk of an increase of the volatile phenols during fermentation. Enzyme addition to the must influenced the aromatic composition of wine in terms of alcohol, volatile acids and esters. Enzyme treatment led to a decrease in protein concentration and a reduction of haze but was ineffective in resolving protein instability. Conclusions Enzyme preparations formulated for aroma release are more prone to induce collateral effects during winemaking. Significance of the Study Our study clearly highlights that enzyme treatment of must may significantly affect the volatile composition of wine. This result can assist the choice of the most suitable enzyme treatment to improve wine composition.

Background and Aims Oxygen (O2) plays a fundamental role in the establishment of wine style and aroma. The effect of O2 treatment during fermentation on the formation of volatile sulfur compounds (VSCs) and the subsequent impact on the sensory profile of wine was investigated. Traditional VSC remediation strategies were also evaluated. Methods and Results Shiraz wines were treated with O2 at several levels during fermentation by sparging rotary fermenters with three types of gas mixtures (40% O2, 21% O2 and N2). The controls were not sparged with any gas and were subsequently subjected to three remedial treatments (aerative racking, early- and late-Cu2+ addition). Wines were analysed for VSCs, fermentation products, concentration of residual metals, as well as their sensory profile. Non-oxygenated ferments and wines contained a higher concentration of VSCs and a lower concentration of fermentation products and differences in the concentration of metals. Volatile sulfur compounds responsive to O2 treatment were identified and a relationship between O2 dose and concentration on VSC formation was demonstrated. The ‘early-Cu2+ addition’ remediation strategy proved the most effective in reducing the impact of ‘reductive’ aromas in non-oxygenated wines. Conclusions The risk of producing a wine with high VSC concentration that negatively impacts wine aroma as well as with a low concentration of fermentation products associated with positive red fruit aromas was decreased by treating wines with O2 during fermentation. Significance of the Study This work specifically examines important aspects of O2 treatment during fermentation in relation to ‘reductive’ aroma characters and the overall impact on the wine's sensory attributes.

Deacidification of wines by malolactic fermentation (MLF) is an essential step in the production of most red, many white and some sparkling base wines. While this secondary fermentation can occur spontaneously, the majority of winemakers, particularly in new world winemaking regions, prefer to minimise the risk of a failed or sluggish MLF by inoculating with a reliable, commercially available starter culture. This review focuses on the scientific literature underpinning growing trends in the application of MLF starters. It considers the literature on co-inoculation regimes, where MLF bacteria are inoculated into a ferment prior to completion of alcoholic fermentation and what benefits this might bring relative to sequential inoculation, when bacteria are added after completion of primary fermentation. It also considers the benefits of the growing trend of using bacteria other than the traditional Oenococcus oeni for induction of MLF, and of developing starter cultures from regional isolates of MLF bacteria to enhance regional identity of wines.

Background and Aims Information regarding the composition and dynamics of microbial communities throughout winemaking is always valuable to control the process and contributes to improving the quality of wine. The aims of this work were to analyse the diversity of lactic acid bacteria (LAB) species present during the spontaneous malolactic fermentation (MLF) of Patagonian red wines, and to isolate and identify native LAB species. Methods and Results Samples of Merlot and Pinot Noir wines were collected during three vintages at one commercial cellar. Through polymerase chain reaction (PCR)-rpoB/denaturing gradient gel electrophoresis (DGGE), it was possible to identify 15 LAB species and through PCR-16S rRNA V3 region/DGGE, 11 LAB species, during spontaneous MLF of these wines, mostly, Oenococcus oeni and Lactobacillus plantarum. These two species were also largely recovered by culture and analysed by the random amplified polymorphic DNA–PCR technique. Isolates of these two LAB species exhibited considerable genotypic heterogeneity. Conclusions The prevalence of O. oeni and Lb. plantarum in all samples suggests that both species are involved in leading the spontaneous MLF of these Patagonian wines. The analysis of the LAB microbiota in wine by PCR-DGGE using two gene regions enabled addition to the number of species that can usually be detected during MLF. Significance of the Study The study offered a more complete knowledge of the LAB community present in wines in a cellar in the Argentine Patagonia during the spontaneous MLF. Our findings are particularly relevant, because they contribute to the possibility of designing an MLF starter composed of native strains of Lb. plantarum and O. oeni.

Background and Aims: Exposure to smoke and uptake of taint imparting phenols in grapes and wines is a significant problem in bushfire-prone regions of Australia and other countries. The effects of smoke exposure on taint occurrence in wines, however, can be variable. This study assessed the influence of cultivar on uptake and accumulation of smoke-borne phenols in grapes and of subsequent processing and winemaking methods on extraction of phenols into wines. Methods and Results: Smoke-exposure experiments were conducted in commercial vineyards of Chardonnay, Merlot and Sauvignon Blanc 14 days after the onset of veraison. At maturity, grapes were harvested for winemaking, which included malolactic fermentation (MLF) for Merlot. Volatile and glycoconjugated phenols were determined in grapes and the resultant wines. All cultivars had a similar concentration of smoke-derived total phenols in their grapes. The apparent extraction of total phenols from grapes into wines, however, differed markedly among the three traditional winemaking methods. Red winemaking (Merlot) with skin contact extracted 88% of total grape phenols, whereas white winemaking either by crushing before pressing (Sauvignon Blanc) or by whole-bunch pressing without crushing (Chardonnay), respectively, released 39 and 18% of total phenols. For Merlot wines, MLF did not affect the extraction of total smoke-derived phenols. Conclusions: Under standardised exposure conditions (duration, intensity and phenology), the three cultivars studied accumulated a similar concentration of total phenols in grapes. The grape-processing and winemaking methods, however, can bring about a fourfold difference in the concentration of total phenols of wines. The smoke-derived phenols extracted from grapes into wine and the distribution of these phenols between the volatile and conjugated pools were not affected by MLF. Significance of the Study: The key findings reported here have the potential to improve decision-making by grapegrowers and winemakers on the effect of cultivar and winemaking practice on potential smoke taint in wine.

Background and Aims Diacetyl accumulation in wine, which has undergone malolactic fermentation by Oenococcus oeni, is often associated with aromatic off-flavours. Characterisation of the diacetyl-related metabolic pathway helps to explain bacterial diacetyl formation during winemaking. The present study describes the time-dependent formation of diacetyl during the vinification process after simultaneous, induced malolactic fermentation by freeze-dried O. oeni on the basis of gene-expression analysis of the citrate- and pyruvate-derived pathways. Methods and Results After simultaneously induced malolactic fermentation in Pinot Blanc by O. oeni, the dynamics of diacetyl formation were compared with citrate consumption and gene-expression of the diacetyl-related metabolic pathways. Diacetyl concentration showed two maxima: the first increase was primarily influenced by the activity of Saccharomyces cerevisiae; however, the second increase was induced only by O. oeni and correlates with bacterial citrate degradation. Expression of the alsS gene showed two significant responses; however, only the second response was affected by the citrate-associated genes maeP and citE. Additionally, alsD and butA2 were found to be continuously underexpressed during the winemaking process. Conclusions Taken together, we suggest that diacetyl accumulation during the vinification process by O. oeni is affected by citrate fermentation. The diacetyl-related alsS gene, however, is also overexpressed independently by other substrates, which may also increase the intracellular pyruvate level resulting in diacetyl formation. Significance of the Study Characterisation of the time-dependent diacetyl accumulation and its degradation during the vinification process is essential for the development of strategies that focus on the suppression of the diacetyl concentration below the sensory threshold.

Background and Aims Glutathione (GSH) is an antioxidant that plays several important roles in wine, including the limitation of browning and of atypical ageing off-flavours and the preservation of important varietal aroma compounds. In finished wines, however, GSH concentration varies significantly. The influence of several oenological factors, including yeast strain, extended lees contact and yeast assimilable nitrogen content, on the concentration of GSH in wine and in a model wine has been investigated. Methods and Results The influence of 20 commercial Saccharomyces cerevisiae wine yeast strains on the GSH content after alcoholic fermentation was evaluated in a chemically defined grape juice (CDGJ). A significant difference was observed between strains, with some strains resulting in a GSH content sevenfold higher than that of other strains. In Sauvignon Blanc grape juice with a range of initial GSH concentration, the concentration of GSH fluctuated during fermentation. After alcoholic fermentation, however, GSH concentration was generally lower than that initially present in the juice; strains that resulted in the highest GSH concentration in the wines prepared from CDGJ did not necessarily display a similar trend in grape juice. Conclusions Yeast strains have a significant impact on the GSH concentration in finished wines, but specific outcomes are dependent on as yet unknown environmental factors. Yeast assimilable nitrogen concentration in a CDGJ did not impact on GSH concentration after fermentation, and GSH concentration generally decreases during ageing irrespective of the yeast strain or the presence of lees. Significance of the Study The data highlight the influence of certain vinification practices on the GSH concentration in wine and suggest strategies to increase the GSH concentration in commercial wines. Such strategies may help to reduce the use of sulfur dioxide by the industry.

Background and Aims Malolactic fermentation (MLF) is a metabolic process promoted in wine by lactic acid bacteria, which can improve wine quality. The MLF, however, must be properly monitored and handled in a timely manner as the bacteria remaining in the wine can produce off-flavours. This work describes the Wireless Sensor Bung (WSB), a system for monitoring remotely wine parameters in barrel and a methodology for monitoring MLF in real time. Methods and Results A wireless system to measure wine deacidification because of MLF was developed. The WSB was calibrated and tested in an experiment in a commercial cellar to assess the system performance in a barrel. The system enabled an exhaustive monitoring of wine deacidification associated with MLF. Conclusions The WSB system was demonstrated to be an effective tool for monitoring remotely MLF in barrel. Significance of the Study The WSB provides timely information on the completion of MLF, so that lactic acid bacteria can be eliminated from wine before they produce undesirable metabolites.

Background and Aims: This study investigated the interactive effects of malolactic bacterial strain, pre-malolactic fermentation (MLF) pH value and wine matrix/style on modulating chemical and sensory impacts of MLF in Cabernet Sauvignon wine. Methods and Results: Malolactic fermentation was conducted in two styles of Cabernet Sauvignon wine, a lighter, fruity style and a more complex style. Each wine was divided into two equal volumes, one was adjusted to pH 3.3 the other to pH 3.7. Each of these wines was further divided into four equal volumes, three of which were inoculated with three different commercial Oenococcus oeni strains and the fourth used as a non-MLF control. Following MLF, all wines were standardised to approximately pH 3.5. The MLF treatments exhibited significant strain- and wine matrix-dependent effects on a diversity of chemical components, including esters, volatile acids and higher alcohols, and on colour. Descriptive sensory analysis also demonstrated significant effects on several sensory properties. Partial least squares analysis revealed a strong correlation between important chemical components and sensory attributes, including overall fruit flavour and dark fruit aroma. Conclusions: The extent and diversity of the impacts of MLF on wine chemical and sensory properties were directly influenced by choice of bacterial strain, pre-MLF pH and wine matrix composition. Relatively harsh conditions (pH 3.3, 14.8% alcohol) protracted the time of MLF and supported greater differences between Oenococcus oeni strains in their modulation of important wine chemical components. Significance of the Study: This study increases knowledge of the significant impacts of MLF on wine chemical composition and associated sensory properties. Specifically, variations in the effect of MLF on wine sensory and chemical properties has been found to arise from the choice of bacterial strain and wine matrix composition/style.

Background and Aims: Phenolic compounds influence colour, flavour and astringency of wines. Technology and grape variety are the main factor affecting the phenolic content of wines. Different multivariate statistical approaches were used to investigate the relationships between the profile of phenolic compounds and grape variety and also the impact of malolactic fermentation (MLF). Methods and Results: A reversed phase liquid chromatography/diode array detection method was used for the analysis of major non-flavonoid phenolic compounds in wines from Trincadeira, Aragonez, Cabernet Sauvignon, Alfrocheiro, Casteao and Touriga Nacional varieties before and after MLF. The impact of MLF and grape variety on phenolic profile was evaluated by principal component analysis (PCA), variation partitioning analysis (VPA) and artificial neural network (ANN). PCA explained 86.5% of the total variance among samples. ANN showed a significant clustering of samples according to grape variety and confirmed that MLF has a minor effect on wine phenolic profile. VPA enabled more information to be extracted from the data by identifying explanatory variables responsible for variability among samples. Conclusions: Compared with PCA and ANN, VPA provides more information concerning the variability on the sample system. Also, grape varieties have a more effective impact on wine low molecular weight phenolic compounds than MLF. Significance of the Study: Each one of the three multivariate statistical approaches showed ways of analysing large chemistry experimental datasets. VPA is a step forward in data analysis, providing more solid and complete assessment of sample system variability, not possible by PCA and ANN.

Background and Aims: This study compared changes in the sensory characteristics of Bobal red wines caused by adding oak chips at different stages of the fermentation process. Methods and Results: The effect of wood, in the form of oak chips, on the sensory characteristics of Bobal wine at different stages of the fermentation process was studied using quantitative descriptive analysis. Bobal control wine was made following traditional winemaking processes, without oak chips. Oak chips were added to the rest of the wines in two dose rates (3 and 6 g/L) at different stages of the winemaking process: for 1 week during alcoholic fermentation, during malolactic fermentation (MLF) and in young, red Bobal wine. The results showed that the Bobal control wine presented a sensory profile characterised by red fruit notes, liquorice, pepper, leather, tobacco and cassis notes. Wines made with the addition of oak chips during alcoholic fermentation presented a similar sensory profile to control wines with some woody (oak) notes. Wines with oak chips added during MLF had a stronger oak character than when used after MLF had completed. Conclusions: It was observed that the use of wood chips gives rise to a different sensorial profile of wines depending at what point of the winemaking process the chips are added. Higher intensities of woody, coconut, vanilla, toasty and toffee descriptors are obtained when a large dose rate of chips is employed. Significance of the Study: All these treatments with oak provide a feasible alternative to traditional winemaking methods as oak chips improve and enhance the sensory profile of Bobal wines.

Background and Aims: Winemakers are constantly searching for new techniques to modulate wine style. Exploiting indigenous yeasts present in grape must is re-emerging as a commercial option in New World wine regions. Wines made with indigenous or 'wild' yeasts are perceived to be more complex by showing a greater diversity of flavours; however, the chemical basis for the flavour characteristics is not yet defined. In order to evaluate techniques for making wine with the 'wild yeast fermentation' character more reliably, it is necessary to define the salient chemical characteristics of such wines. Methods and Results: Pairs of Chardonnay wines were prepared from the same must and subjected to similar fermentation conditions in the wineries of origin, except for the mode of inoculation. Reference wines were made by inoculation with a Saccharomyces cerevisiae starter culture, whereas companion wines were allowed to undergo fermentation with the indigenous microflora. Of all wine chemicals analysed, only yeast-derived volatile fermentation products showed significant differences between the yeast treatments. Conclusions: Inoculated wines were associated with the esters ethyl hexanoate and 3-methylbutyl acetate and formed a clear cluster by principal component analysis. By comparison with inoculated wines, 'wild' yeast fermented wines showed high variability in volatile compounds that contribute to wine aroma, with higher concentrations of 2-methylpropanol, 2-methylbutanoic acid, ethyl 2-methylpropanoate, ethyl decanoate and ethyl dodecanoate potentially being sensorially important. Significance of the Study: This study shows that yeast-derived volatile fermentation products are a key difference between inoculated and uninoculated ferments and provides a chemical basis for the 'wild yeast fermentation' character.

Red wine was made from the same batch of Cabernet Sauvignon grapes but with two different yeasts, Saccharomyces cerevisiae (SC) or Saccharomyces bayanus (SB). The colour properties and pigment profiles of SC and SB wines were compared at 8 days and 387 days after yeast inoculation. Grape anthocyanin concentration was found to be lower in SB wines than in SC wines at day 8 and 387, but SB wine exhibited greater wine colour density, a* and b*. The anthocyanin concentration did not correlate with wine colour density. The levels of pigmented polymers and SO2 non-bleachable pigments were found to be higher in SB wine at day 387, demonstrating that the formation of stable pyranoanthocyanins and pigmented polymers was enhanced by SB yeast. Electrospray mass spectrometry was used for the compositional analysis of pyranoanthocyanins, and direct and ethyl-bridged linked condensation dimers of malvidin 3-O-glucoside (MvG) with flavanols (catechin or epicatechin). The data demonstrated that the formation of acetaldehyde-mediated pigments was enhanced by the use of the SB yeast. The compositional analysis suggested that the differences in colour properties and pigment profiles of SC and SB wines were largely due to the greater production of acetaldehyde-mediated pigments by the use of SB yeast.

Sparkling wine produced by the traditional methode champenoise requires a second in-bottle alcoholic fermentation of a base wine, leading to the sparkling wine. This second fermentation is followed by prolonged ageing in contact with yeast cells (lees). The autolysis of yeast occurs during the ageing of sparkling wines. During this process, the yeast releases different compounds that modify the organoleptic properties of the wine. The ageing period is required to give these wines their roundness and characteristic aroma and flavour. Autolysis products also influence the foaming properties of sparkling wine. Yeast autolysis is characterised by the hydrolysis of intracellular biopolymers by yeast enzymes activated after cell death. This results in the release of low molecular weight products. This article reviews the recent advances in understanding the yeast autolysis mechanism, the factors affecting autolysis, the nature of the released compounds and their effects on sparkling wine quality.

The effect of high sugar concentrations or the presence of copper residues on fermentation efficiency and volatile acid (VA) production were examined for selected commercial wine yeast strains. Fermentation rate as measured by accumulated mass loss appeared to be least affected by high sugar concentrations during the first few days of fermentation for strains VIN13, WE14, N96 and RJ11. It was, however, only fermentations using VIN13, WE372, N96 and L2056 that contained less than 11 g/L fructose after 21 days. VIN13 and RJ11 produced the lowest VA in the 21°, 25° and 28°Brix musts. Fermentation efficiency of six strains was also studied in must containing 0.25 mM Cu2+. Results indicated that strains NT50, Collection Cepage Cabernet (CC) and D80 were not significantly affected, whereas VIN13, NT112 and RJ11 contained significantly more glucose and fructose after fermentation in the must containing copper. Copper addition also increased VA production, with strains RJ11 and NT50 being the most affected.

The ability of different wine yeast (Saccharomyces cerevisiae) to inhibit malolactic bacteria (Oenococcus oeni) and the influence of nitrogen were studied using a synthetic grape juice. Malolactic fermentation was induced in fermenting synthetic grape juice or synthetic wines inoculated with different commercial strains of S. cerevisiae. O. oeni was generally inhibited in wines that contained higher concentrations of total SO2 although many yeast strains only inhibited the bacteria during fermentation under high nitrogen conditions. Yeast produced higher amounts of SO2 during fermentation under high nitrogen conditions suggesting that nitrogen affected the malolactic fermentation by influencing yeast SO2 production. However, the production of SO2 by yeast did not always account for the inhibition of O. oeni, suggesting the presence of other inhibitory mechanisms.

This review discusses the impacts of nitrogen addition in the vineyard and winery, and establishes the effects that nitrogen has on grape berry and wine composition and the sensory attributes of wine. Nitrogen is the most abundant soil-derived macronutrient in a grapevine, and plays a major role in many of the biological functions and processes of both grapevine and fermentative microorganisms. Manipulation of grapevine nitrogen nutrition has the potential to influence quality components in the grape and, ultimately, the wine. In addition, fermentation kinetics and formation of flavour-active metabolites are also affected by the nitrogen status of the must, which can be further manipulated by addition of nitrogen in the winery. The only consistent effect of nitrogen application in the vineyard on grape berry quality components is an increase in the concentration of the major nitrogenous compounds, such as total nitrogen, total amino acids, arginine, proline and ammonium, and consequently yeast-assimilable nitrogen (YAN). Both the form and amount of YAN have significant implications for wine quality. Low must YAN leads to low yeast populations and poor fermentation vigour, increased risk of sluggish/stuck/slow fermentations, increased production of undesirable thiols (e.g. hydrogen sulfide) and higher alcohols, and low production of esters and long chain volatile fatty acids. High must YAN leads to increased biomass and higher maximum heat output due to greater fermentation vigour, and increased formation of ethyl acetate, acetic acid and volatile acidity. Increased concentrations of haze-causing proteins, urea and ethyl carbamate and biogenic amines are also associated with high YAN musts. The risk of microbial instability, potential taint from Botrytis-infected fruit and possibly atypical ageing character is also increased. Intermediate must YAN favours the best balance between desirable and undesirable chemical and sensory wine attributes. ‘Macro tuning’, of berry nitrogen status can be achieved in the vineyard, given genetic constraints, but the final ‘micro tuning’ can be more readily achieved in the winery by the use of nitrogen supplements, such as diammonium phosphate (DAP) and the choice of fermentation conditions. This point highlights the need to monitor nitrogen not only in the vineyard but also in the must immediately before fermentation, so that appropriate additions can be made when required. Overall, optimisation of vineyard and fermentation nitrogen can contribute to quality factors in wine and hence affect its value. However, a better understanding of the effect of nitrogen on grape secondary metabolites and different types of nitrogen sources on yeast flavour metabolism and wine sensory properties is still required.

Wine is a highly complex mixture of compounds which largely define its appearance, aroma, flavour and mouth-feel properties. The compounds responsible for those attributes have been derived in turn from three major sources, viz. grapes, microbes and, when used, wood (most commonly, oak). The grape-derived compounds provide varietal distinction in addition to giving wine its basic structure. Thus, the floral monoterpenes largely define Muscat-related wines and the fruity volatile thiols define Sauvignon-related wines; the grape acids and tannins, together with alcohol, contribute the palate and mouth-feel properties. Yeast fermentation of sugars not only produces ethanol and carbon dioxide but a range of minor but sensorially important volatile metabolites which gives wine its vinous character. These volatile metabolites, which comprise esters, higher alcohols, carbonyls, volatile fatty acids and sulfur compounds, are derived from sugar and amino acid metabolism. The malolactic fermentation, when needed, not only provides deacidification, but can enhance the flavour profile. The aroma and flavour profile of wine is the result of an almost infinite number of variations in production, whether in the vineyard or the winery. In addition to the obvious, such as the grapes selected, the winemaker employs a variety of techniques and tools to produce wines with specific flavour profiles. One of these tools is the choice of microorganism to conduct fermentation. During alcoholic fermentation, the wine yeast Saccharomyces cerevisiae brings forth the major changes between grape must and wine: modifying aroma, flavour, mouth-feel, colour and chemical complexity. The wine bacterium Oenococcus oeni adds its contribution to wines that undergo malolactic fermentation. Thus flavour-active yeasts and bacterial strains can produce desirable sensory results by helping to extract compounds from the solids in grape must, by modifying grape-derived molecules and by producing flavour-active metabolites. This article reviews some of the most important flavour compounds found in wine, and their microbiological origin.

Malolactic fermentation (MLF) is the bacterial-driven decarboxylation of L-malic acid to L-lactic acid and carbon dioxide, and brings about deacidification, flavour modifications and microbial stability of wine. Pasteur first described the presence of ‘bacteria’ in wine nearly one-hundred-and-fifty years ago and the subsequent elucidation of the bacterial-driven malolactic reaction was described about fifty years later. Over the following years the occurrence of MLF became apparent in wines worldwide, and eventually Oenococcus oeni was identified as the principal organism involved in the process. O. oeni is remarkable in its ability to tolerate the nutritionally poor and challenging, harsh wine environment; however, it can be a difficult and sometimes unreliable organism to work with in the winery. A greater knowledge of its biology would undoubtedly facilitate the development of strains and practices, with improved performance outcomes. We already know a considerable amount about the biochemistry and physiology of O. oeni, and ironically, although we know little about its genetics, its genome has been sequenced. With this groundwork in place and molecular biology techniques at our disposal we are poised to increase our knowledge and understanding of this organism enormously.

Lysozyme is an enzyme with muramidase activity which can lyse Gram-positive bacteria, including wine lactic acid bacteria. This enzyme provides a practical method for delaying or preventing the growth of Oenococcus oeni and consequently the onset of malolactic fermentation during the vinification of red and white wines. This paper reports the impact of lysozyme on the chemical and sensorial properties of commercially vinified red (Cabernet Sauvignon and Shiraz) and white (Riesling) wines. The addition of lysozyme to these wines had little or no effect on the content of alcohol, free and total sulfur dioxide and titratable acidity, and pH value. The lysozyme retained 75-80% activity in the Riesling wine after six months, however no detectable activity remained in the Cabernet Sauvignon and Shiraz wines after two days. Upon addition of lysozyme to both of the red wines, a rapid initial decrease (up to 17%) in red wine colour density and phenolic content occurred in association with the formation of a light precipitate. The reduction in red wine colour was also noted by the sensory panel. When added to the Riesling wine, lysozyme did not cause an increase in browning over the six month storage time, but did induce heat instability (haze), suggesting that white wines may require protein stabilisation following treatment with lysozyme. Sensory assessment by triangle difference testing revealed that, during the six month storage period, treatment with lysozyme did not cause important changes to either the aroma or palate of the red and white wines tested.

The effect of the interaction between pH (2.9-3.5), alcohol concentration (12.5-14.5% v/v) and wine matrix (Chardonnay, Grenache blend and Cabernet Sauvignon) on malolactic fermentation (MLF) induced by two commercially available directly inoculated Oenococcus oeni starter cultures was determined. Wine matrix had greatest impact on the rate of MLF, followed by pH and alcohol; the matrix effect was significantly modified by pH and bacterial culture. The rate of L-malic acid degradation by starter culture A was 3.3-61.3% in the Grenache blend and 3.3-4.5% in the Chardonnay wine compared to the Cabernet Sauvignon wine, for the pH/alcohol ranges 2.9/14.5 to 3.5/12.5. L-malic acid degradation rate by starter culture B was lower but less affected by wine matrix. The rate of L-malic acid degradation was closely related to culture viability, with a high rate of L-malic acid degradation being associated with bacterial growth and vice versa. Treatment of two wines in which MLF had previously failed to complete with various combinations of nutrients, carbon fining and heating, restored MLF, suggesting that a growth-limiting nutrient was absent and/or a toxic substance not related to pH, alcohol or SO2 concentration was present. This work demonstrated that the wine matrix may affect culture viability more significantly than either pH or alcohol concentration. A high rate of MLF was shown to depend on factors permitting bacterial growth, however, the two strains studied responded differently to the wine matrix, suggesting that malolactic bacteria vary in tolerance to various stresses in the wine environment.

The present study was undertaken to evaluate the influence of medium pH and inoculation time on the growth and malolactic activity of an Oenococcus oeni culture. Samples of a commercial white grape juice adjusted to pH 3.2, 3.4 or 3.6, and inoculated with Saccharomyces cerevisiae strain CY3079, were inoculated with a malolactic culture (Oenococcus oeni strain 31) at the beginning, middle, and end of alcoholic fermentation. The results obtained from this single case study show that it is possible to inoculate the bacterial culture at the three different times during alcoholic fermentation without slowing down or stopping alcoholic fermentation or causing failure of MLF. However, pH and timing of bacterial inoculation were critical to how rapidly MLF starts. At pH 3.2 a lowering of bacterial viability was observed, but a more important reduction was recorded at all tested pH levels when the bacteria were inoculated halfway through alcoholic fermentation. When inoculation was carried out at the end of alcoholic fermentation, the presence of yeast seemed to favour bacterial viability and activity and bacteria performed MLF even in difficult conditions such as pH values around 3. In all wines malolactic fermentation was accompanied by total degradation of malic and citric acids and production of L-lactic acid, D-lactic and acetic acids.

Intraspecific variation of Oenococcus oeni, the preferred lactic acid bacteria species for inducing malolactic fermentation in wine, was studied using the randomly amplified polymorphic DNA (RAPD) strain fingerprinting technique. Ten of fifteen isolates of O. oeni from Australian wineries situated in different wine regions could be distinguished by the RAPD technique. Strains of O. oeni which originated from the same winery were either indistinguishable or closely related to each other. Six different commercially available O. oeni strains could be differentiated with the four RAPD primers used and their genetic similarity determined. Analysis of O. oeni present in wines from a single source of fruit (Cabernet Sauvignon, vintage 2002) that underwent spontaneous malolactic fermentation revealed wide genetic variation amongst the isolates. Each fermentor contained several different O. oeni strains, which were present throughout alcoholic and malolactic fermentation. These data highlight the sensitivity of RAPDs when suitable primers are applied to O. oeni of unknown genetic origin, thus enabling O. oeni strains with desirable sensory and fermentation properties to be genetically analysed.

Metabolic interaction of wine yeast and malolactic (ML) bacteria may stimulate or inhibit malolactic fermentation (MLF). To study this interaction in the absence of extrinsic grape-derived and processing factors, a standardised methodology using a chemically defined medium was developed to reproducibly characterise the intrinsic metabolic interaction between any wine yeast and bacteria pair. The methodology can be adapted to study the modifying effects of extrinsic factors on yeast-bacteria interaction. The methodology involves measuring the growth characteristics of the chosen ML bacterium in the wine produced by the yeast of interest under standardised conditions. The methodology was applied to four Saccharomyces cerevisiae wine yeasts which have been characterised in terms of growth response of an Oenococcus oeni strain. Three yeasts gave compatible interactions, in which the level of bacterial growth stimulation was variable; the fourth yeast inhibited bacterial growth and produced a high concentration of sulfite. To test for the possibility that the bacterial growth response was inhibited by the depletion of nutrients by the fermentation yeast, supplementation of the wines with essential nutrients gave a strain-dependent growth response. A treatment designed to remove or inactivate yeast-derived inhibitory factors was also investigated. This treatment partly alleviated bacterial inhibition caused by the sulfite-producing yeast strain. To verify that the standardised methodology relates to grape juice and wine, the synthetic grape juice medium was replaced with a Chardonnay juice of comparable composition prepared from microbiologically sound grapes; similar results were obtained, suggesting that the grape-derived extrinsic factors in this case did not appreciably modify the intrinsic yeast-bacteria metabolic interaction.

During alcoholic fermentation the formation of vitisin A occurred mainly in the period between 20% and 85% glucose utilisation, when the concentrations of the precursors, malvidin-3-glucoside and pyruvic acid were at a maximum. The maximum rate of vitisin A synthesis of 11 mg/Lh occurred when 57% of glucose had been utilised and 440 mg/L malvidin-3-glucoside, 114 mg/L pyruvic acid and 3.1 mg/L measured oxygen were present. During maturation, production of vitisin A appeared to be linked with the availability of a suitable oxidant. Synthesis of vitisin A continued for approximately 6 months of maturation in air-tight bottles while the precursors were available. In wines that underwent malolactic fermentation, the malolactic bacteria consumed pyruvic acid and thereby limited the production of vitisin A. At the end of a 12 month maturation period, the concentration of vitisin A declined in wines made without any malolactic fermentation, even though malvidin-3-glucoside and pyruvic acid were still present. This indicated that there was a shortage of a suitable oxidant required to complete the reaction.

An important flavour compound associated with malolactic fermentation (MLF) is the volatile di-ketone, diacetyl (2,3-butanedione), which is considered as the major contributor to buttery flavour in wines. Commercial Chardonnay, Cabernet Sauvignon, Merlot and Shiraz wines from eight vintages (1992- 1999), 20 viticultural regions and 28 wine producers were analysed for diacetyl and also the buttery aroma intensity was rated by a trained sensory panel. The wines examined were found to have a wide range of diacetyl concentration, i.e. 0.3 to 0.6 mg/L (mean value 0.4 mg/L) for the 24 Chardonnay wines, and 0.3 to 2.5 mg/L (mean value 1.1 mg/L) for the 43 red wines. The buttery aroma, while significantly correlated with diacetyl concentration in the Chardonnay wines (P < 0.01), was not predicted very well by the regression equation, but the prediction was improved when the free sulfur dioxide concentration of the wines was taken into account (adjusted R2 = 0.43, P < 0.01). There was a weak, although statistically significant, correlation between diacetyl concentration and the buttery rating for the red wine data set, when the sulfur dioxide concentration was taken into account (adjusted R2 = 0.12, P < 0.05). This weak correlation may have been, in part, due to the generally low concentration of diacetyl in relation to the published aroma threshold in red wine. It was also noted that there was a substantial decrease in diacetyl concentration for a subset of red wines, but not for white wines, of various vintages cellared for three years at 15°C.

The efficacy of lysozyme against indigenous lactic acid bacteria (LAB) and four inoculated spoilage LAB cultures was investigated in laboratory scale Chardonnay winemaking trials (at pH 3.8). These LAB cultures included Lactobacillus kunkeei, Lactobacillus brevis, Pediococcus parvulus, and Pediococcus damnosus. Three concentrations of lysozyme were used: 0, 125 and 250 mg/L. Alcoholic fermentation of the grape juice was carried out at 20±0.5oC using Saccharomyces cerevisiae. Lysozyme did not have any negative impact on yeast growth and sugar reduction. This enzyme was found to be very effective in inhibiting the growth of all four LAB cultures investigated. Under the given experimental conditions, as high as an 8 log cell reduction was obtained for some of the strains. The acetic acid production by L. brevis and L. kunkeei was significantly reduced in the treatments with 125 and 250 mg/L lysozyme added (P < 0.01). The effect of lysozyme on the cells of the LAB cultures was examined under a scanning electron microscope. It is evident that lysozyme had a detrimental impact on the cells of these cultures. Based on these observations, it is concluded that lysozyme may be a useful tool for winemakers to control the growth of spoilage LAB and to reduce the production of volatile acids. The addition of lysozyme may also prevent the increase of volatile acidity during stuck/sluggish alcoholic fermentation. This tool is particularly useful in high pH wines where SO2 is less effective.

Interactions between the yeast strain used for primary oenological fermentation and the bacterium used to conduct subsequent malolactic fermentation were studied under model winemaking conditions. A commercial Saccharomyces cerevisiae wine yeast (strains, EC 1118, AWRI 835 and CY-3079) was grown in a defined medium whose composition approximated grape juice. Fermentations by all strains reached dryness, and retained a cell viability of greater than 90% upon completion of fermentation. Highest total viable cell number and percentage of viable cells were recorded for EC 1118. A sur lie ageing of the fermented medium over a 12 week period revealed a bi-phasic decay of culture viability for all strains. Thus 99% of cells had died within 2 weeks post-fermentation. Viabilities were then stable for the subsequent 4-6 week period before a second decline phase ensued and ended in either a minimal (ca 100 CFU/mL, EC 1118) or no viable cells being detected at 12 weeks of ageing. The growth response of an Oenococcus oeni inoculum to yeast culture supernatants, previously aged for up to 12 weeks in the presence or absence of yeast lees, was evaluated in a bio-assay. In this way, yeast strains could be designated as being either inhibitory, neutral or stimulatory to the growth of O. oeni (strain Lc5p). Inhibition by supernatants of strain EC 1118 was evident, but found to be reduced by ageing the supernatant (with or without lees). Conversely, longer ageing on yeast lees increased the magnitude of the stimulatory response in O. oeni (strain Lc5p) to the supernatant from the wine yeast (strain CY-3079).

Descriptive sensory profiles of Uruguayan Tannat wines following malolactic fermentation (MLF) with Oenococcus oeni were compared with control wines where MLF was prevented. MLF led to a significant decrease (P < 0.05) in secondary descriptors such as 'berry fruit' and 'fresh vegetative', as well as a decrease in related tertiary descriptors such as 'blackcurrant', 'apricot', 'cut green grass' and 'green pepper'. The main differences in the chemical composition of Tannat wine aromas following MLF were (1) an increase of the lactates (mainly ethyl lactate) over the sensory threshold; (2) a significant decrease in ethyl esters and acetates, depending on the bacteria strain utilised (the most significant change was found in hexyl acetate), and (3) a small increase of 4-vinylguaiacol and 4-vinylphenol with one of the MLF strains (this change was below the threshold of human perception). We also observed an increase of compounds related to a-ketoglutarate metabolism, namely: ethyl 4-hidroxibutanoate and g{butyrolactone. Differences in chemical composition due to MLF are discussed in relation to variation in sensory profiles.