Ethylene-responsive VviERF003 modulates glycosylated monoterpenoid synthesis by upregulating VviGT14 in grapes.

Terpenoids are important contributors to the aroma of grapes and wines. Grapes contain terpenoids in both volatile free form and non-volatile glycosidic form, with the latter being more abundant. Glycosylated terpenoids are deemed as latent aromatic potentials for their essential role in adding to the flowery and fruity bouquet of wines. However, the transcriptional regulatory mechanism underlying glycosylated terpenoid biosynthesis remains poorly understood. Our prior study identified an AP2/ERF transcription factor, VviERF003, through DNA pull-down screening using the promoter of terpenoid glycosyltransferase VviGT14 gene. This study demonstrated that both genes were co-expressed and synchronized with the accumulation of glycosylated monoterpenoids during grape maturation. VviERF003 can bind to the VviGT14 promoter and promote its activity according to yeast one-hybrid and dual-luciferase assays. VviERF003 upregulated VviGT14 expression in vivo, leading to increased production of glycosylated monoterpenoids based on the evidence from overexpression or RNA interference in leaves, berry skins, and calli of grapes, as well as tomato fruits. Additionally, VviERF003 and VviGT14 expressions and glycosylated monoterpenoid levels were induced by ethylene in grapes. The findings suggest that VviERF003 is ethylene-responsive and stimulates glycosylated monoterpenoid biosynthesis through upregulating VviGT14 expression.

Identification of SNP loci and candidate genes genetically controlling norisoprenoids in grape berry based on genome-wide association study.

Obtaining new grapevine varieties with unique aromas has been a long-standing goal of breeders. Norisoprenoids are of particular interest to wine producers and researchers, as these compounds are responsible for the important varietal aromas in wine, characterized by a complex floral and fruity smell, and are likely present in all grape varieties. However, the single-nucleotide polymorphism (SNP) loci and candidate genes genetically controlling the norisoprenoid content in grape berry remain unknown. To this end, in this study, we investigated 13 norisoprenoid traits across two years in an F1 population consisting of 149 individuals from a hybrid of Vitis vinifera L. cv. Muscat Alexandria and V. vinifera L. cv. Christmas Rose. Based on 568,953 SNP markers, genome-wide association analysis revealed that 27 candidate SNP loci belonging to 18 genes were significantly associated with the concentrations of norisoprenoid components in grape berry. Among them, 13 SNPs were confirmed in a grapevine germplasm population comprising 97 varieties, including two non-synonymous mutations SNPs within the VvDXS1 and VvGGPPS genes, respectively in the isoprenoid metabolic pathway. Genotype analysis showed that the grapevine individuals with the heterozygous genotype C/T at chr5:2987350 of VvGGPPS accumulated higher average levels of 6-methyl-5-hepten-2-one and ß-cyclocitral than those with the homozygous genotype C/C. Furthermore, VvGGPPS was highly expressed in individuals with high norisoprenoids concentrations. Transient overexpression of VvGGPPS in the leaves of Vitis quinquangularis and tobacco resulted in an increase in norisoprenoid concentrations. These findings indicate the importance of VvGGPPS in the genetic control of norisoprenoids in grape berries, serving as a potential molecular breeding target for aroma.

Grape polysaccharides: compositional changes in grapes and wines, possible effects on wine organoleptic properties, and practical control during winemaking.

Polysaccharides present in grapes interact with wine sensory-active compounds (polyphenols and volatile compounds) via different mechanisms and can affect wine organoleptic qualities such as astringency, color and aroma. Studies on the role that grape polysaccharides play in wines are reviewed in this paper. First, the composition of grape polysaccharides and their changes during grape ripening, winemaking and aging are introduced. Second, different interaction mechanisms of grape polysaccharides and wine sensory-active compounds (flavanols, anthocyanins and volatiles) are introduced, and the possible effects on wine astringency, color and aroma caused by these interactions are illustrated. Finally, the control of the grape polysaccharide content in practice is discussed, including classical winemaking methods (applying different maceration enzymes, temperature control, co-fermentation, blending), modern vinification technologies (pulsed electric field, ultrasound treatment), and the development of new grape polysaccharide products.

Red wine coloration: A review of pigmented molecules, reactions, and applications.

Color is one of the most distinctive qualities of red wine. Despite new knowledge in the field of pigment identification, copigmentation, and oxidation being forthcoming, there is still a large gap between the fundamental research and practical winemaking outcomes. A state-of-art review from these two aspects is, therefore, necessary. This review first introduces updated knowledge about the primary pigments in wine, with emphasis on their physicochemical properties. Then, the mechanisms of copigmentation and oxidation are elucidated in detail, along with their relative contributions to wine color. Finally, the practical effects of copigmentation and micro-oxygenation (MOX) in winemaking are summarized and discussed. In general, wine coloration is ultimately determined by the anthocyanin flavylium cation, which is greatly influenced by wine pH. In young red wine, grape-derived anthocyanins and nonanthocyanin polyphenols (as copigments) are the foundation for wine coloration. During aging and storage, anthocyanin derivatives are formed via various chemical reactions, where moderate oxidation plays a vital role, whereas copigmentation constantly decreases. The essence of wine color evolution relates to the changes of physicochemical properties of primary pigments in wine, where the hydration equilibrium gradually diminishes. In practice, the effects of copigment addition and MOX during real vinification can be viewed as somewhat controversial, considering that many studies showed different effects on wine color and pigment concentration. Universal features can be summarized but some phenomena still remain unclear and deserve further exploration.

Characterization of odor-active compounds in the head, heart, and tail fractions of freshly distilled spirit from Spine grape (Vitis davidii Foex) wine by gas chromatography-olfactometry and gas chromatography-mass spectrometry.

Differences in key odor-active volatile compounds among the head, heart, and tail fractions of freshly distilled spirits from Spine grape (Vitis davidii Foex) wine were identified for the first time by gas chromatography-olfactometry and gas chromatography-mass spectrometry. Results from aroma extract dilution analysis (AEDA) showed that there were 34, 45, and 37 odor-active compounds in the head, heart and tail fractions, respectively. Besides, 20, 22, and 17 quantified compounds, respectively, showed odor activity values (OAVs) > 1. The head fraction was characterized by fruity, fusel/solvent notes owing to higher concentrations of higher alcohols and esters, while the tail fraction had more intense smoky/animal, sweaty/fatty attributes due to higher concentrations of volatile phenols and fatty acids. Finally, the heart fraction was characterized by ethyl octanoate, ethyl hexanoate, ethyl 3-phenylpropanoate, ethyl cinnamate, isoamyl alcohol, guaiacol, 4-ethylguaiacol, 4-vinylguaiacol, 2,3-butanedione, and (E)-ß-damascenone. Furthermore, observation of the distillation progress indicated that different volatiles with various boiling points and solubilities followed diverse distillation patterns: concentrations of most esters, higher alcohols, terpenes and C13-norisoprenoids decreased, while concentrations of volatile phenols, fatty acids and some aromatic compounds increased during distillation. As a result, their final concentrations in the three distillate fractions varied significantly.

Influence of attenuated reflected solar radiation from the vineyard floor on volatile compounds in Cabernet Sauvignon grapes and wines of the north foot of Mt. Tianshan.

In this study, fruit-zone microclimate was modified by three treatments, including inter-row mulch (M), the combination of leaf removal applied at the onset of veraison and inter-row mulch (MLR-BV), and the combination of leaf removal applied at complete veraison and inter-row mulch (MLR-EV), in a semi-arid climate in three consecutive years (2015-2017). M decreased fruit-zone reflected solar radiation from vineyard floor and low temperature (10-20 °C) duration, whereas it increased soil temperature and high temperature (> 30 °C) duration. MLR-BV and MLR-EV increased fruit-zone incident photosynthetically active radiation while decreased the duration of 20-25 °C compared to M. Notably, M significantly decreased grape total norisoprenoid concentrations in 2015-2017, and total terpenoid concentrations in 2015-2016. Applying leaf removal applied at the onset of veraison could compensate the decreases of total norisoprenoids and terpenoids caused by M when two treatments were applied together. Besides, M significantly increased grape total C6/C9 compound concentrations, besides, (Z)-3-hexen-1-ol concentrations were significantly higher in grapes of M than those of MLR-BV in 2015-2017. Light exposure and high temperature duration after veraison had strong positive correlations with total norisoprenoids and terpenoids, besides, low temperature duration was positively correlated with total norisoprenoids. In addition, light exposure after veraison had strong negative correlations with total C6/C9 compounds. With respect to the volatile compounds in wines, M significantly decreased the concentrations of isopentanol and ethyl acetate, and the concentrations of ethyl cinnamate, phenylacetaldehyde, phenylethyl alcohol and 3-methylthio-1-propanol were significantly lower in MLR-BV and MLR-EV than in M. The outcome of this study can assist winegrowers to properly adjust vineyard managements to optimize the concentrations of desired volatile compounds in grapes and wines.

Characterization of Transcriptional Expression and Regulation of Carotenoid Cleavage Dioxygenase 4b in Grapes.

Norisoprenoids are important aromatic volatiles contributing to the pleasant floral/fruity odor in grapes and wine. They are produced from carotenoids through the cleavage of carotenoid cleavage dioxygenases (CCDs). However, the underlying mechanisms regulating VvCCD expression remain poorly understood. In this study, we showed that VvCCD4b expression was positively correlated with the accumulation of ß-damascenone, ß-ionone, 6-methyl-5-hepten-2-one, geranylacetone, dihydroedulan I, and total norisoprenoids in developing grapes in two vintages from two regions. VvCCD4b was found to be principally expressed in flowers, mature leaves, and berries. Abscisic acid strongly induced the expression of this gene. Additionally, the present study preliminarily indicated that the activity of the VvCCD4b promoter was dropped under 37°C treatment and also responded to the illumination change. VvCCD4b was expressed in parallel with VvMADS4 in developing grape berries. The latter is a MADS family transcription factor and nucleus-localized protein that was captured by yeast one-hybrid. A dual-luciferase reporter assay in tobacco leaves revealed that VvMADS4 downregulated the activity of the VvCCD4b promoter. VvMADS4 overexpression in grape calli and Vitis quinquangularis Rehd. leaves repressed the VvCCD4b expression. In summary, this work demonstrates that VvCCD4b expression is positively correlated with the accumulation of norisoprenoids, and VvMADS4 is a potential negative regulator of VvCCD4b. Our results provide a new perspective for understanding the regulation of VvCCD4b expression and norisoprenoid accumulation in grapes.

Blending strategies for wine color modification â : Color improvement by blending wines of different phenolic profiles testified under extreme oxygen exposures.

Limited oxygenation and over-oxidation experiments were designed to compare the phenolic and chromatic characters of base wines Cabernet Franc (CF), Cabernet Sauvignon (CS), and their counterparts that blended with modifier wines Marselan (MA) and Petit Verdot (PV). In both limited oxygenation and over-oxidation conditions, all blend wines generally contained higher C*ab, a* and Red%, and lower hab, b* and Yellow% than their base wine counterparts, because MA contributed flavonols (copigments) and anthocyanins, and PV contributed flavanols (anthocyanin derived pigments precursors). Chromatic changes that can be perceived by human eye (ΔE*ab) in CF based blend wines were more obvious than that of CS based blend wines, which indicate that base wine with lower phenolic concentrations and weak phenolic profiles (CF) might be more prone to be chromatically modified than base wine with higher phenolic concentrations and distinct phenolic profiles (CS). Chemical influences of different blending strategies on anthocyanin derivatives' formations were depending on phenolic profiles of the modifier wines and base wines, and also on the oxygen exposure. The results suggest that the chromatic improvement of base wines could be realized by blending modifier wines under different oxygen exposures.

Characterization and differentiation of key odor-active compounds of 'Beibinghong' icewine and dry wine by gas chromatography-olfactometry and aroma reconstitution.

Freezing-thawing events contribute to the unique aroma profile of icewines. Differences in key odor-active volatile compounds between 'Beibinghong' (Vitis amurensis × V. vinifera) icewines and dry wines were investigated by gas chromatography-olfactometry and gas chromatography-mass spectrometry. Acceptable agreement between the olfactometric and quantitative results was obtained. 'Beibinghong' icewine was characterized by high concentrations of volatile phenols, lactones, (E)-ß-damascenone, and phenylacetaldehyde, which were associated with on-vine freezing-thawing events in grape. Low concentrations of higher alcohol acetates and ethyl esters of fatty acids were attributed to hyperosmotic stress during fermentation. The overall aroma of icewine could be mimicked by reconstitution containing 44 identified volatiles. Partial least squares regression analysis demonstrated that the concentrations of these volatile compounds determined the distinct sensory profiles of icewines, which have higher intensities of honey/sweet, smoky, caramel, dried fruit, apricot/peach, and floral aromas, and lower intensities of fresh fruity and herbaceous notes in comparison with dry wines.

Changes in monosaccharides, organic acids and amino acids during Cabernet Sauvignon wine ageing based on a simultaneous analysis using gas chromatography-mass spectrometry.

BACKGROUND: Monosaccharides, organic acids and amino acids are the important flavour-related components in wines. The aim of this article is to develop and validate a method that could simultaneously analyse these compounds in wine based on silylation derivatisation and gas chromatography-mass spectrometry (GC-MS), and apply this method to the investigation of the changes of these compounds and speculate upon their related influences on Cabernet Sauvignon wine flavour during wine ageing. This work presented a new approach for wine analysis and provided more information concerning red wine ageing. RESULTS: This method could simultaneously quantitatively analyse 2 monosaccharides, 8 organic acids and 13 amino acids in wine. A validation experiment showed good linearity, sensitivity, reproducibility and recovery. Multiple derivatives of five amino acids have been found but their effects on quantitative analysis were negligible, except for methionine. The evolution pattern of each category was different, and we speculated that the corresponding mechanisms involving microorganism activities, physical interactions and chemical reactions had a great correlation with red wine flavours during ageing. CONCLUSION: Simultaneously quantitative analysis of monosaccharides, organic acids and amino acids in wine was feasible and reliable and this method has extensive application prospects. © 2017 Society of Chemical Industry.

The Influence of Prefermentative Addition of Gallic Acid on the Phenolic Composition and Chromatic Characteristics of Cabernet Sauvignon Wines.

In this study, the prefermentative addition of gallic acid in Cabernet Sauvignon red winemaking was performed. The influence of gallic acid addition on wine phenolic composition, the ratio of copigmentation, and the color parameters were monitored throughout the winemaking process. The results showed that the prefermentative addition of gallic acid enhanced the extraction of total anthocyanins and the copigmentation effect, producing wines with more darkness, redness, yellowness, and saturation. Moreover, the addition of gallic acid contributed to the concentration of total phenolic acids. However, it had a negative effect on the concentrations of flavonols and flavan-3-ols in the final wines. Thus, the prefermentative addition of gallic acid at appropriate levels might be a promising enological technology to obtain wines with high color quality and aging potential.

Light response and potential interacting proteins of a grape flavonoid 3'-hydroxylase gene promoter.

Flavonoid 3'-hydroxylase (F3'H), a member of cytochrome P450 protein family, introduces B-ring hydroxyl group in the 3' position of the flavonoid. In this study, the cDNA sequence of a F3'H gene (VviF3'H), which contains an open reading frame of 1530 bp encoding a polypeptide of 509 amino acids, was cloned and characterized from Vitis vinifera L. cv. Cabernet Sauvignon. VviF3'H showed high homology to known F3'H genes, especially F3'Hs from the V. vinifera reference genome (Pinot Noir) and lotus. Expression profiling analysis using real-time PCR revealed that VviF3'H was ubiquitously expressed in all tested tissues including berries, leaves, flowers, roots, stems and tendrils, suggesting its important physiological role in plant growth and development. Moreover, the transcript level of VviF3'H gene in grape berries was relatively higher at early developmental stages and gradually decreased during véraison, and then increased in the mature phase. In addition, the promoter of VviF3'H was isolated by using TAIL-PCR. Yeast one-hybrid screening of the Cabernet Sauvignon cDNA library and subsequent in vivo/vitro validations revealed the interaction between VviF3'H promoter and several transcription factors, including members of HD-Zip, NAC, MYB and EIN families. A transcriptional regulation mechanism of VviF3'H expression is proposed for the first time.

A review of polyphenolics in oak woods.

Polyphenolics, which are ubiquitous in plants, currently are among the most studied phytochemicals because of their perceptible chemical properties and antioxidant activity. Oak barrels and their alternatives, which are widely used in winemaking nowadays, contribute polyphenolics to wines and are thought to play crucial roles in the development of wines during aging. This study summarizes the detailed information of polyphenolics in oak woods and their products by examining their structures and discussing their chemical reactions during wine aging. This paper evaluates the most recent developments in polyphenolic chemistry by summarizing their extraction, separation, and their identification by the use of chromatographic and spectral techniques. In addition, this paper also introduces polyphenol bioactive ingredients in other plant foods.

Association between modification of phenolic profiling and development of wine color during alcohol fermentation.

To solve the problem of wine color instability in western China, different additives (the maceration enzymes Vinozym G and Ex-color, yeasts VR5 and Red Star, and commercial tannins) were added during alcoholic fermentation of Syrah (Vitis vinifera L.). The phenolic profile and color characteristics of wine were examined using high performance liquid chromatography mass spectrometry and CIELAB, respectively. The results showed that the combination of the enzyme Ex-color with the Red Star yeast eased the release of non-anthocyanins from grape berries into wine, whereas the use of enzyme Vinozym G and VR5 yeast enhanced the concentration of anthocyanins and achieved a higher red hue (a* value) and a lower yellow hue (b* value) in the wine. The addition of commercial tannins greatly promoted the level of gallic acid in the wine and led to a relatively higher concentration of anthocyanins. Partial least-squares regression analysis was used to find out the major phenolics, which were in close relation with color parameters; principal component analysis was used to evaluate the contribution of different winemaking techniques to wine color. The combination of these 2 analytic methods indicated that Vinozym G and VR5 yeast together with commercial tannins should be an appropriate combination to enhance the stability of wine color during alcohol fermentation, which was related to a significant increase in cyanidin-3-O-(6-O-acetyl)-glucoside, cyanidin-3-O-(6-O-coumaryl)-glucoside, trans-peonidin-3-O-(6-O-coumaryl)-glucoside, trans-malvidin-3-O-(6-O-coumaryl)-glucoside, and malvidin-3-O-(6-O-acetyl)-glucoside-pyruvic acid, all of which played an important role in stabilizing wine color.

Biosynthesis and genetic regulation of proanthocyanidins in plants.

Proanthocyanidins (PAs), also known as condensed tannins, are a group of polyphenolic secondary metabolites synthesized in plants as oligomers or polymers of flavan-3-ol units via the flavonoid pathway. Due to their structural complexity and varied composition, only in the recent years has the study on the biosynthesis and regulation of PAs in plants taken off, although some details of the synthetic mechanism remain unclear. This paper aims to summarize the status of research on the structures of PAs in plants, the genes encoding key enzymes of biosynthetic pathway, the transport factors, the transcriptional regulation of PA biosynthesis and the genetic manipulation of PAs. The problems of this field were also discussed, including the nature of the final "enzyme" which catalyzes the polymerization reaction of PAs and the possible mechanism of how the elementary units of flavanols are assembled in vivo.