Enhancing grapevine breeding efficiency through genomic prediction and selection index.

Grapevine (Vitis vinifera) breeding reaches a critical point. New cultivars are released every year with resistance to powdery and downy mildews. However, the traditional process remains time-consuming, taking 20-25 years, and demands the evaluation of new traits to enhance grapevine adaptation to climate change. Until now, the selection process has relied on phenotypic data and a limited number of molecular markers for simple genetic traits such as resistance to pathogens, without a clearly defined ideotype, and was carried out on a large scale. To accelerate the breeding process and address these challenges, we investigated the use of genomic prediction, a methodology using molecular markers to predict genotypic values. In our study, we focused on 2 existing grapevine breeding programs: Rosé wine and Cognac production. In these programs, several families were created through crosses of emblematic and interspecific resistant varieties to powdery and downy mildews. Thirty traits were evaluated for each program, using 2 genomic prediction methods: Genomic Best Linear Unbiased Predictor and Least Absolute Shrinkage Selection Operator. The results revealed substantial variability in predictive abilities across traits, ranging from 0 to 0.9. These discrepancies could be attributed to factors such as trait heritability and trait characteristics. Moreover, we explored the potential of across-population genomic prediction by leveraging other grapevine populations as training sets. Integrating genomic prediction allowed us to identify superior individuals for each program, using multivariate selection index method. The ideotype for each breeding program was defined collaboratively with representatives from the wine-growing sector.

Diffusion of phenolic compounds during a model maceration in winemaking: role of flesh and seeds.

BACKGROUND: During red winemaking, diffusion of phenolic compounds from the grape berry cells into the liquid phase occurs simultaneously with the adsorption of the same compounds onto the pulp. In previous studies, we quantified the proportions of polyphenols diffusing from the skins and then assessed the amounts that can be fixed by the pulp. In this work, we added the impact of seeds, also present during vinification, by carrying out macerations in a model medium with the following berry compartments: skins, seeds, skins + seeds, skins + seeds + pulp. RESULTS: Interestingly, the seeds alone released a rather high amount of polyphenols. As soon as they were in the presence of cell walls of skin/flesh, and/or anthocyanins, the concentration of seed tannins in the solution dropped dramatically, due to a combined effect of adsorption and/or precipitation and/or chemical reactions. The pulp certainly adsorbed tannins, but they also tended to shift the extraction equilibria, and it seems that more tannins could be extracted from skins and seeds when pulp was present. Polyphenol amounts extracted in model systems with skins + seeds + pulp were close to what was extracted in microvinification. CONCLUSION: These model experiments reflect relatively well extraction during microvinification experiments and highlight the respective impact of the grape berry's different compartments in the wine's final phenolic composition as well as some of the mechanisms involved. © 2022 Society of Chemical Industry.

Focus on the relationships between the cell wall composition in the extraction of anthocyanins and tannins from grape berries.

Concentrations of anthocyanins and tannins after extraction from berries in wines and from skin macerations in model solutions have been studied for two grape varieties, two maturation levels and two vintages berries. Characterization of the cell wall polysaccharides has also been performed, the classical method based on the analysis of the neutral sugars after depolymerization being completed by a comprehensive microarray polymer profiling (CoMPP). Extraction was lower in model solutions than in wines, with the same ranking: non acylated anthocyanins> tannins > p-coumaroylated anthocyanins. The polysaccharidic composition suggested a role of homogalacturonans, rhamnogalacturonans and extensins in the extraction process. A global explanation of the interactions between anthocyanins, tannins and polysaccharides is proposed.

Multi-method study of the impact of fermentation on the polyphenol composition and color of Grenache, Cinsault, and Syrah rosé wines.

Rosé wines show large color diversity, due to different phenolic pigment compositions. However, the mechanisms responsible for such diversity are poorly understood. The present work aimed at investigating the impact of fermentation on the color and composition of rosé wines made from Grenache, Cinsault, and Syrah grapes. Targeted MS analysis showed large varietal differences in must and wine compositions, with higher concentrations of anthocyanins and flavanols in Syrah. UV-visible spectrophotometry and size exclusion chromatography data indicated that Grenache and Cinsault musts contained oligomeric pigments derived from hydroxycinnamic acids and flavanols which were mostly lost during fermentation due to adsorption on lees. Syrah must color was mainly due to anthocyanins which were partly converted to derived pigments through reactions with yeast metabolites with limited color drop during fermentation. This work highlighted the impact of must composition, reflecting varietal characteristics, on changes occurring during fermentation and consequently wine color.

Elucidating the Color of Rosé Wines Using Polyphenol-Targeted Metabolomics.

The color of rosé wines is extremely diverse and a key element in their marketing. It is due to the presence of anthocyanins and of additional pigments derived from them and from other wine constituents. To explore the pigment composition and determine its links with color, 268 commercial rosé wines were analysed. The concentration of 125 polyphenolic compounds was determined by a targeted metabolomics approach using ultra high-performance liquid chromatography coupled to triple quadrupole mass spectrometry (UHPLC-QqQ-MS) analysis in the Multiple Reaction Monitoring (MRM) mode and the color characterised by spectrophotometry and CieLab parameters. Chemometrics analysis of the composition and color data showed that although color intensity is primarily determined by polyphenol extraction (especially anthocyanins and flavanols) from the grapes, different color styles correspond to different pigment compositions. The salmon shade of light rosé wines is mostly due to pyranoanthocyanin pigments, resulting from reactions of anthocyanins with phenolic acids and pyruvic acid, a yeast metabolite. Redness of intermediate color wines is related to anthocyanins and carboxypoyranoanthocyanins and that of dark rosé wines to products of anthocyanin reactions with flavanols while yellowness of these wines is associated to oxidation.

Evolution of Flavanol Glycosides during Red Grape Fermentation.

Monomeric and dimeric flavanol glycosides were quantified by UHPLC-MRM in Syrah (SYR) and Grenache (GRE) grapes and in their corresponding wines for the first time. Quantities were extremely variable depending on grape tissue (seeds or skins) and during fermentation. Overall, 22 monomeric and dimeric mono- and diglycosides were determined with concentrations ranging from 0.7 nanograms to 0.700 micrograms per gram of grape tissue, and 0 to 60 micrograms per liter for wines. The evolution of the glycosides' composition during winemaking suggests that almost all these compounds originate in the grapes themselves and display different extraction kinetics during winemaking. One isomer of the monomeric (epi) flavanol monoglycosides seemed to be biosynthesized by yeasts during wine fermentation. The sharp decrease in concentration of some isomers at the late stages of fermentation or after pressing suggests that some grape glycosidase activities convert these compounds into non-glycosylated flavanols.

Innovative analysis of 3-mercaptohexan-1-ol, 3-mercaptohexylacetate and their corresponding disulfides in wine by stable isotope dilution assay and nano-liquid chromatography tandem mass spectrometry.

Both 3-mercaptohexan-1-ol (3MH) and 3-mercaptohexyl acetate (3MHA) were largely studied for the last 20 years due to their pleasant olfactory notes conferred to wine. Until now, many analytical methods focused only on the free forms of both 3MH and 3MHA in wine that provided partial information in the wine aroma evolution. Our study proposes new analytical measurements which allow quantification of both free and disulfide forms of 3MH and 3MHA to better understand the redox phenomenon occurring in wines and further, to orientate wine aroma evolution. Free thiols were analyzed by an original method based on maleimide derivatization allowing in-situ disulfide reduction followed by SIDA-LC-MS/MS analyses exhibiting excellent performances. Indeed, the accuracy ranged from 95 to 110% in three different wine matrices and the repeatability and intermediate reproducibility were inferior to 15% (RSD measurements). Our method exhibited very low limits of detection, which are below to 0.5ng/L and inferior to the perception thresholds of both compounds. Then, this method was applied to three different wines exposed to several oxidative conditions. On the one hand, it was demonstrated that copper sulfate treatment firstly destroyed the total amount of free 3MH to the benefit of thioether and disulfides compounds, with proportions that could be slightly modified by glutathione addition. On the other hand, oxygenation of wines resulted in partial free 3MH destruction to the benefit of thioether compounds. We proposed for the first time an innovative analysis that gives a complete picture of wine aroma, which can be really useful to winemakers to manage wine aroma evolution and to take advantage of the disulfide reservoir.

Effect of pulsed electric field treatment during cold maceration and alcoholic fermentation on major red wine qualitative and quantitative parameters.

This work studies the effect of pulsed electric field (PEF) treatment at moderate and high field strengths (E=0.8kV/cm & 5kV/cm) prior and during alcoholic fermentation (AF) of red grapes on improving different parameters of pre-treated extracts: pH, °Brix, colour intensity (CI), total polyphenols content (TPI) of Cabernet Sauvignon red wine. Similar trends were observed for treating grapes using moderate and high electric field strength on the enhancement of CI and TPI of the wine after AF. The application of PEF using moderate strengths at different times during cold maceration (CM) (0, 2 and 4days) was more efficient for treatment during CM. The treatment during AF showed lower extraction rate compared to treating during CM and prior to AF. Our results clearly show that the best time for applying the PEF-treatment through the red fermentation is during the CM step.

Changes in polyphenol profiles and color composition of freshly fermented model wine due to pulsed electric field, enzymes and thermovinification pretreatments.

This work compares the effects of three pretreatments techniques: pulsed electric fields (PEFs), enzymes treatment (ET) and thermovinification (TV) on the improving of extraction of main phenolic compounds, color characteristics (L(∗)a(∗)b(∗)), and composition (copigmentation, non-discolored pigments) of freshly fermented model wine from Cabernet Sauvignon variety. The pretreatments produced differences in the wines, with the color of the freshly fermented model wine obtained from PEF and TV pretreated musts being the most different with an increase of 56% and 62%, respectively, compared to the control, while the color only increased by 22% for ET. At the end of the alcoholic fermentation, the contents of anthocyanins for all the pretreatments were not statistically different. However, for the content of total phenolics and total flavonols, PEF and TV were statistically different, but ET was not statistically different. The contents of flavonols in musts pretreated by PEF and TV were significantly higher comparing to the control. An increase of 48% and 97% was noted respectively, and only 4% for ET. A similar result was observed for the total phenolics with an increase by 18% and 32% respectively for PEF and TV, and only 3% for ET comparing to the control. The results suggest that the higher intensity and the difference of color composition between the control and pretreated freshly fermented model wines were not related only to a higher content of residual native polyphenols in these freshly fermented model wines. Other phenomena such as copigmentation and formation of derived pigments may be favored by these pretreatments.

Effect of macerating enzymes on the oligosaccharide profiles of Merlot red wines.

Commercial pectinase preparations are applied in winemaking to improve wine processing and final quality. These preparations contain pectolytic enzyme activities such as polygalacturonases, pectin esterases, pectin lyases, and rhamnogalacturonases. These enzymes modify the polysaccharide and oligosaccharide composition of wines. The influence of various commercial enzyme preparations on wine oligosaccharide composition was studied, on Merlot wines from the Bordeaux area. Wine oligosaccharides were isolated by high-resolution size-exclusion chromatography on a Superdex-30 HR column. The glycosyl residue and glycosyl linkage compositions of the oligosaccharide fractions obtained were determined. The MS spectra of the Merlot oligosaccharide fractions from control and enzyme-treated wines were recorded on an AccuTOF mass spectrometer equipped with an electrospray ionization (ESI) source and a time-of-flight (TOF) mass analyzer. Oligosaccharides in the control wines were partly methylated homogalacturonans, corresponding to smooth regions of pectins, whereas those of the enzyme-treated wines were mostly rhamnogalacturonan-like structures linked with neutral lateral chains, arising from the hairy regions. The enzyme preparations used thus cleaved the rhamnogalacturonan backbone of the hairy zones and demethylated and hydrolyzed the smooth regions. Besides, different structures were detected, depending on the enzyme preparation used, indicating that they contained rhamnogalacturonase activities with different specificities. The oligosaccharide profiles can serve as a marker of enzymatic treatments.

Modeling procyanidin self-association processes and understanding their micellar organization: a study by diffusion NMR and molecular mechanics.

The colloidal behavior of eight synthetic procyanidins (three monomers, four dimers, and a trimer) has been investigated in water or in a winelike medium using DOSY NMR spectroscopy and molecular dynamics simulations. Different behavior was observed for monomers and oligomers. Monomers self-associate with a high affinity constant (37-53 M(-1)) to form micelles at low cmc (critical micelle concentration) values (1-5 g.L(-1)). These micelles undergo a time-dependent coalescence process to form hazes and precipitates. As for dimers and the trimer, self-association also occurs but with a lower affinity (approximately 6 M(-1)) and at higher cmc values (10-20 g.L(-1)) to form small micelles (<5 nm) that remain stable throughout the experiment. The presence of 10% ethanol does not significantly affect the self-association constant for monomers and oligomers but increases their cmc values by approximately 50% and decreases the micelle size by a factor 2. However, the presence of 20 mM NaCl appears to negate the effect of ethanol. This study helps to clarify the role of procyanidin monomers versus oligomers in wine turbidity and demonstrates that procyanidin oligomers are fully available to interact with saliva proteins.

Structural and conformational analysis of two native procyanidin trimers.

The structure and conformation of two native procyanidin trimers in water have been determined using 2D NMR and molecular mechanics. The results show the existence of four rotameric forms, one of which is predominant (60 to 80%). These four rotamers are shown to be in slow to intermediate exchange on the NMR timescale. Both trimers, whose structures vary owing to a different substitution of one carbon atom, adopt conformations in which stacking between different phenolic rings is favored.

A 3D structural and conformational study of procyanidin dimers in water and hydro-alcoholic media as viewed by NMR and molecular modeling.

The three-dimensional structures of 5 procyanidin dimers have been determined in a hydro-alcoholic medium and in water using 2D NMR and molecular mechanics. They are made from monomers of catechin (CAT) and epicatechin (EPI)-B1: EPI-CAT, B2: EPI-EPI, B3: CAT-CAT, B4: CAT-EPI and B2g: EPI-EPI-3-O-gallate. These tannins exist in two conformations that are in slow exchange in the NMR timescale (s), one is compact and the other extended. The compact form is found to dominate (76-98%) when the dimer is made of at least one CAT monomer (B1, B3, B4). Both forms are found in even proportions only in the case of procyanidin B2. The latter tannin can be converted into a dominant compact form when the lower EPI unit is galloylated. The finding of a predominant compact form for procyanidin dimers is discussed in relation with tannin-saliva protein interactions that are of importance for the wine-tasting/making processes.