Anthocyanins and flavan-3-ols from grapes and wines of Vitis vinifera cv. Cesanese d'Affile

The objective of the present study was to evaluate the amount of some potential health-promoting phenols in the grape of Vitis vinifera cv. Cesanese d'Affile and in wines made from these grapes. The analyses were performed using HPLC/DAD/MS. The accumulation of anthocyanins in the skin and flavan-3-ols in the seed was determined at different stages of ripening of the grape (i.e. green, veraison, middle stage of ripening, and complete ripening). Thirteen anthocyanins were identified in the skin at all stages of ripening, except the green stage. With regard to flavan-3-ols, (+)-catechin, (-)-epicatechin, and (-)-epicatechin gallate were detected in all of the seed samples. The highest (+)-catechin content was found in the seeds of the green grape (2 mg g(-1) DW), whereas in the seeds from the completely ripe grape the content was more than ten times lower. The highest catechin content in the seed was correlated with the lowest anthocyanin content in the skin. The wines produced in the years 2004 and 2005 showed, at wavelengths of 520 and 280 nm, almost identical quali-quantitative chromatographic profiles, with high concentrations of anthocyanin 3-O-glucosides, low concentrations of acylated anthocyanins, and trace amounts of (+)-catechin and (-)-epicatechin.     

Polyphenols from Vitis vinifera Seeds

The chemical composition of polyphenols from grape seeds was studied. It has been found that they contain four catechins and two proanthocyanidins. The probable structure of the proanthocyanidins was proposed on the basis of physicochemical investigations.     

Monoterpenic and norisoprenoidic glycoconjugates of Vitis vinifera L. cv. Melon B. as precursors of odorants in Muscadet wines.

The volatile monoterpenic and norisoprenoidic compounds released by glycosidase enzyme hydrolysis of C18 reversed-phase isolates from the juice of Vitis vinifera L. cv. Melon B. have been qualitatively and quantitatively determined using GC-MS and GC-FID. The components analyzed were broadly similar to those previously reported for other varieties but the level of bound p-menth-1-en-7,8-diol was higher in this cultivar. Then the monoterpenic and norisoprenoidic volatiles released from the same glycosidic extracts under mild acid conditions, mimicking wine aging conditions, have been analyzed using GC-Olfactometry and GC-MS. The most odorous compounds detected were p-cymene, terpinen-4-ol, cis- and trans-vitispiranes, 1,6,6-trimethyl-1,2-dihydronaphtalene (TDN), beta-damascenone and riesling acetal. To assess their potential levels in corresponding wines after ageing, most of these odorants were generated by harsh acid hydrolysis from the precursors extracts and quantitatively determined using SPME and GC-MS/MS. For the development and application of this analysis, the odorants not commercialy available were synthesized. The total amounts of norisoprenoidic odorants generated by acid hydrolysis of the glycosidic extracts were shown to be proportional to the total amounts of these precursors.     

Redox Behavior of Anthocyanins Present in Vitis vinifera L.

Voltammetric techniques were employed to study the electrochemical behavior of several anthocyanins. The redox behavior of anthocyanins with the same basic structure, the influence of glycosylation on the redox behavior of anthocyanins derived from different anthocyanidins, and the influence of methoxylation were investigated. The anthocyanins used in this study were malvidin‐3‐O‐glucoside chloride, malvidin‐3,5‐di‐O‐glucoside chloride, cyanidin‐3‐O‐glucoside chloride, cyanidin‐3,5‐di‐O‐glucoside chloride, peonidin‐3‐O‐glucoside chloride, delphinidin‐3‐O‐glucoside chloride and the anthocyanidin petunidin chloride, all of them present in Vitis vinifera L. All hydroxyl groups of the anthocyanins can be electrochemically oxidized and the anthocyanins studied revealed a complex and pH dependent oxidation process, with the occurrence of adsorption and of oxidation products blocking the electrode surface.     

Preparative isolation of polyphenolic compounds from Vitis vinifera by centrifugal partition chromatography

This study deals with a centrifugal partition chromatography developed for the separation of phenolic compounds from Vitis vinifera. EtOAc grape seed extracts were separated using the solvent system hexane-ethyl acetate-ethanol-water (1:8:2:7; v/v) in two fractions: one containing about 75% of flavanol monomers (catechin and epicatechin) corresponding to 18% of crude extract and another fraction B-type dimers (22% of crude extract). From the stalk extracts, we could separate stilbenoid compounds (resveratrol and its oligomers; 12% of crude extract) which were eluted in less than 30 min from flavanols (which required a few hours of additional elution). Using the same solvent system but in different ratios (4:5:3:3; v/v), we isolated the trans-resveratrol (7@1000; 90% purity).     

Application of Elicitors in Two Ripening Periods of Vitis vinifera L. cv Monastrell: Influence on Anthocyanin Concentration of Grapes and Wines

In recent years, it has been demonstrated that the application of elicitors such as methyl-jasmonate (MeJ) and benzothiadiazole (BTH) to wine grapes can increase their phenolic and aromatic compounds if they are treated at the beginning of ripening (veraison). However, the veraison period is short, and it is not always possible to apply the treatments in a few days. Therefore, it would be of great interest to optimize the moment of elicitor application or extend the treatment period. The aim of this paper was to analyze during two consecutive years (2016–2017) the foliar application of MeJ, BTH, and a combination of both, during two different ripening periods of Monastrell grapes (veraison and mid-ripening), and determine the more appropriate moment to increase the concentration of anthocyanins. To carry out this aim, analysis of anthocyanins by HPLC in grapes and wines was mainly performed. The most suitable period for the application of MeJ, BTH, and MeJ + BTH was at mid-ripening, since the grapes showed a greater accumulation of anthocyanins at harvest. However, the MeJ + BTH treatment applied during veraison also obtained similar results, which would allow extending the application period if necessary. However, the increase in the anthocyanin content of grapes was not reflected in all the wines, which may have been due to reinforcement of the skin cell wall as a result of the application of elicitors. Further analysis is needed to improve the maceration process of the Monastrell grapes and the extraction of the anthocyanins that were increased by the treatments applied in the vineyard.     

Aqueous extract from Vitis vinifera tendrils is able to enrich keratinocyte antioxidant defences

An aqueous extract of V. vinifera L. tendrils was evaluated for its ability to enrich the antioxidant capacity of cultured cells. The long-time antioxidant capability of the extract was measured by in vitro chemical methods, and its influence on reduced glutathione levels and plasma membrane oxido reductase activity was determined in cultured human keratinocytes (NCTC 2544). Keratinocytes are cells normally exposed to oxidative stress, and for this reason adequately equipped with antioxidant defences. However, it has long been suggested that exogenous antioxidants may play an important role in minimizing the adverse effects of oxidative stress on skin.We demonstrated that V. vinifera tendril aqueous extract was able to increase, in a time- and dose-dependent manner, the reduced glutathione concentration and activity of trans plasma membrane oxido reductase as an indirect evaluation of the intracellular redox status of the cells demonstrating a relevant antioxidant activity of this phytocomplex.     

Oxidative Transformation of Dihydroflavonols and Flavan-3-ols by Anthocyanidin Synthase from Vitis vinifera

Twelve polyphenols from three distinct families (dihydroflavonols, flavan-3-ols, and flavanones) were studied as potential substrates of anthocyanidin synthase from Vitis vinifera (VvANS). Only flavan-3-ols of (2R,3S) configuration having either a catechol or gallol group on ring B are accepted as substrates. Only dihydroflavonols of (2R,3R) configuration are accepted as substrates, but a catechol or gallol group is not mandatory. Flavanones are not substrates of VvANS. HPLC and MS/MS analyses of the enzymatic products showed that the VvANS-catalyzed oxidative transformation of (+)-dihydroflavonols, such as dihydroquercetin, dihydrokaempferol and dihydromyricetin, leads only to the corresponding flavonols. Among the flavan-3-ols recognized as substrates, (+)-gallocatechin was only transformed into delphinidin by VvANS, whereas (+)-catechin was transformed into three products, including two major products that were an ascorbate–cyanidin adduct and a dimer of oxidized catechin, and a minor product that was cyanidin. Data from real-time MS monitoring of the enzymatic transformation of (+)-catechin suggest that its products are all derived from the initial C₃-hydroxylation intermediate, i.e., a 3,3-gem-diol, and their most likely formation mechanism is discussed.     

High‐performance liquid chromatography/electrospray ionization tandem mass spectrometric investigation of stilbenoids in cell cultures of Vitis vinifera L., cv. Malvasia

Cell suspension cultures of Vitis vinifera L., particularly after elicitation, have shown the ability to biosynthesize several stilbenoids. In this work the application of specific tandem mass spectrometry (MS/MS) experiments and the integration with data from UV‐vis spectra, available only for the main compounds of the extract, allowed the detection of over 20 different stilbenoids. Up to eight monoglicosides, both monomers and dimers, a high number of dimer forms (up to 17), belonging to the group of viniferins and to their oxidized derivatives, were identified in the extract of Malvasia cell cultures. Moreover, the selectivity and sensitivity of the method allowed detection also of a few cis derivatives (up to 3) present in very low concentrations and presumably produced by the light exposure during treatment of the sample. To the best of the authors' knowledge, there are no literature data on MS/MS experiments targeted to simultaneously study monomeric and dimeric stilbenoids, both as free and glycosilated forms, in a complex mixture from V. vinifera cell suspension cultures. Copyright © 2010 John Wiley & Sons, Ltd.     

Profiling monoterpenol glycoconjugation in Vitis vinifera L. cv. Muscat of Alexandria using a novel putative compound database approach,high resolution mass spectrometry and collision induced dissociation fragmentation analysis

In this work we present a novel approach for the identification of plant metabolites using ultrahigh performance liquid chromatography coupled to accurate mass time-of-flight mass spectrometry. The workflow involves developing an in-house compound database consisting of exact masses of previously identified as well as putative compounds. The database is used to screen accurate mass spectrometry (MS) data to identify possible compound matches. Subsequent tandem MS data is acquired for possible matches and used for structural elucidation. The methodology is applied to profile monoterpene glycosides in Vitis vinifera cv. Muscat of Alexandria grape berries over three developmental stages. Monoterpenes are a subclass of terpenes, the largest class of plant secondary metabolites, and are found in two major forms in the plant, “bound” to one or more sugar moieties or “free” of said sugar moieties. In the free form, monoterpenes are noted for their fragrance and play important roles in plant defense and as attractants for pollinators. However, glycoconjugation renders these compounds odorless, and it is this form that the plant uses for monoterpene storage. In order to gain insight into monoterpene biochemistry and their fate in the plant an analysis of intact glycosides is essential. Eighteen monoterpene glycosides were identified including a monoterpene trisaccharide glycoside, which is tentatively identified here for this first time in any plant. Additionally, while previous studies have identified monoterpene malonylated glucosides in other grapevine tissue, we tentatively identify them for the first time in grape berries. This analytical approach can be readily applied to other plants and the workflow approach can also be used for other classes of compounds. This approach, in general, provides researchers with data to support the identification of putative compounds, which is especially useful when no standard is available.     

Supercritical fluid extraction of resveratrol from grape skin of Vitis vinifera and determination by HPLC

Supercritical fluid extraction of resveratrol from grape skin of Vitis vinifera was studied. Extraction variables such pressure, modifier concentration (ethanol), and extraction time were optimised. Final extraction conditions were: 40 degrees C, 150 bar, 7.5% ethanol and extraction time 15 min. Extraction recovery and precision (variation coefficient between 0.2 and 1.0%) were calculated. The resveratrol content in the ethanolic extract was determined by HPLC with UV detection at 306 nm. Acetic acid-methanol-water was used as the mobile phase, and C-18 and C-8 columns were tested, instrumental parameters were optimised, and analytical parameters were calculated (lineal interval 0-75 mg l(-1), detection limit 0.1 mg l(-1), sensitivity 125530 mg(-1) mg l(-1), coefficient variation 0.8-1.6%). Six different varieties of grape skin, from the same geographical area and representative of the wine elaboration, were analysed.     

Ecophysiological and phytochemical response to ozone of wine grape cultivars of Vitis vinifera L.

Vitis vinifera sensitivity to tropospheric ozone (O₃) has been evidenced in several studies. In this work, physiological and metabolic effects of O₃ on two wine cultivars of V. vinifera (i.e. Maturano and San Giuseppe) have been studied. Moreover, chlorogenic acid (CGA) production, in consideration of its importance in the biosynthetic pathway of polyphenols and as antioxidant, has been investigated. Maturano cultivar resulted more sensitive to O₃, as evidenced by the gas exchange reduction at the early stage of treatment, and by the increase in Ci/Ca and the decoupling of net photosynthesis and the stomatal conductance at the end of the treatment. Unexpectedly, O₃ did not activate stilbene production. Ozone induced an early CGA decrease, significantly more consistent in cv. Maturano, and an increase after 8 days, more consistent in cv. S. Giuseppe. These results suggest that CGA could be considered a biochemical marker of O₃-induced stress in V. vinifera.     

High Temperature Affects Photosynthetic and Molecular Processes in Field‐Cultivated Vitis vinifera L. × Vitis labrusca L.

High‐temperature stress markedly influences grape growth and development. However, how high‐temperature stress response differs between controlled and field‐cultivated grape is poorly understood. In this study, the effects of heat treatment on grapevines were studied for changes in photosystem II (PSII) activity and expression levels of heat‐responsive genes and heat shock protein HSP21. July 31st, 2015 was considered as the post high‐temperature treatment (“42°C”; temperatures above 40°C for a period of time each day ranging from 1–7 h) under field cultivation in our experiment. The recovery of chlorophyll fluorescence indicators and the increasing expression of heat‐responsive genes and the heat shock protein HSP21 suggested the development of heat tolerance in the form of acclimation in grape. Changes in various parameters of photosynthetic pigment fluorescence and of the electron transport chain (Fv/Fm, PI_ABS, W_k, RC_QA, Φ_Po, and Φ_Eo) between “42°C” and the 45°C treatment demonstrated that the donor side, reaction center, and acceptor side of PSII were influenced by a critical high temperature. Furthermore, the difference between the two cultivation conditions studied was attributed to other environmental factors and inherent tree vigor.     

Differentiation of Vitis vinifera varieties by MALDI-MS analysis of the grape seed proteins

Until now the study of pathogenic related proteins in grape juice and wine, performed by ESI-MS, LC/ESI-MS, and MALDI/MS, has been proposed for differentiation of varieties. In fact, chitinases and thaumatin-like proteins persist through the vinification process and cause hazes and sediments in bottled wines. An additional instrument, potentially suitable for the grape varieties differentiation, has been developed by MALDI/MS for the grape seed protein analysis. The hydrosoluble protein profiles of seeds extract from three different Vitis vinifera grape (red and white) varieties were analyzed and compared. In order to evaluate the environmental conditions and harvest effects, the seed protein profiles of one grape variety from different locations and harvests were studied.     

Study on the Chemical Composition and the Biological Activities of Vitis vinifera Stem Extracts

Vitis vinifera (V. vinifera) is a herbaceous plant, cultivated worldwide and known for its biological benefits. The aim of this study is the investigation of the chemical composition as well as the determination of the biological potential of different grape stem extracts obtained by maceration and accelerated solvent extraction (ASE). The HPLC analysis of the tested extracts led to the identification of 28 compounds of which 17 were identified for the first time in grape plants, in addition to seven revealed in the stem part for the first time. Twenty-nine volatile molecules have been detected by GC-MS in the grape stem part; among them seven were identified for the first time in the grape plant. For the biological analysis, the ethyl acetate extract (EtOAc) obtained by maceration showed a significant potential regarding antioxidant activity (IC₅₀ = 42.5 µg/mL), anti-Alzheimer (IC₅₀ = 14.1 µg/mL), antidiabetic (IC₅₀ = 13.4 µg/mL), cytotoxic with HCT-116 (IC₅₀ = 12.5 µg/mL), and anti-inflammatory (IC₅₀ = 26.6 µg/mL) activities, as well as showing the highest polyphenol content (207.9 mg GAE/g DW).     

Impact of the Acetaldehyde-Mediated Condensation on the Phenolic Composition and Antioxidant Activity of Vitis vinifera L. Cv. Merlot Wine

Acetaldehyde is a critical reactant on modifying the phenolic profile during red wine aging, suggesting that the acetaldehyde-mediated condensation can be responsible for the variation of antioxidant activity during the aging of this beverage. The present study employs exogenous acetaldehyde at six levels of treatment (7.86 ± 0.10–259.02 ± 4.95 mg/L) before the bottle aging of Merlot wines to encourage phenolic modification. Acetaldehyde and antioxidant activity of wine were evaluated at 0, 15, 30, 45, 60 and 75 days of storage, while monomeric and polymeric phenolics were analyzed at 0, 30 and 75 days of storage. The loss of acetaldehyde was fitted to a first-order reaction model, the rate constant (k) demonstrated that different chemical reaction happened in wines containing a different initial acetaldehyde. The disappearance of monomeric phenolics and the formation of polymeric phenolics induced by acetaldehyde could be divided into two phases, the antioxidant activity of wine did not alter significantly in the first phase, although most monomeric phenolics vanished, but the second phase would dramatically reduce the antioxidant activity of wine. Furthermore, a higher level of acetaldehyde could shorten the reaction time of the first phase. These results indicate that careful vinification handling aiming at controlling the acetaldehyde allows one to maintain prolonged biological activity during wine aging.     

Separation and identification of highly fluorescent compounds derived from trans-resveratrol in the leaves of Vitis vinifera infected by Plasmopara viticola

A method for identification of highly fluorescent compounds in vine leaves infected by Plasmopara viticola was developed using reversed phase liquid chromatography with simultaneous diode array and fluorometric detection. Fluorescent compounds were extracted from leaves with a methanol-water mixture (70:30). Separation by HPLC was performed using a C(18) column and gradient elution with water-acetonitrile mixtures (20-80% of acetonitrile). The main unknown fluorescent compound was identified by line spectral comparison with a standard obtained by UV photoisomerization of trans-resveratrol glucoside, and its structure was confirmed by liquid chromatography-mass spectrometry. Identification and structural elucidation of the fluorescent compound in the leaves of Vitis vinifera allows early detection of Plasmopara viticola invasion.     

Comparison of anthocyanins present in grapes of Vitis vinifera L. varieties and interspecific hybrids grown in the Czech Republic

Anthocyanins present in the grapes of nine grapevine (Vitis vinifera L.) varieties and ten interspecific hybrids were compared. Total anthocyanin levels were determined in fresh grapes and the values ranged from 0.50 g kg^?1 to 4.99 g kg^?1. A total of twenty-two different anthocyanins were identified. Malvidin 3-glucoside was the most abundant anthocyanin, with concentrations ranging from 30 % to 64 % of the totals observed. The ratio of acetylated to coumaroylated malvidin and peonidin derivatives ranged from 0.09 (variety: Regent) to 1.34 (variety: Cerason), depending on the variety. Diglucosylated anthocyanins were not found in particular interspecific hybrids such as the teinturier XIV 26–56, Laurot, Merlan, and Nativa.     

Impact of Climatic Conditions on the Resveratrol Concentration in Blend of Vitis vinifera L. cvs. Barbera and Croatina Grape Wines

The aim of this work was to investigate the effect of meteorological conditions on resveratrol concentration of red wines produced in Piacenza viticultural region (Italy). In this regard, six representative estates producing Colli Piacentini Gutturnio DOC (a blend of V. vinifera L. cvs. Barbera and Croatina) vintage wines were analysed for trans- and cis-resveratrol over an 8-year period (1998–2005). Grapes were taken from the same vineyard in each estate by using the same enological practices over the entire investigated period. The meteorological conditions corresponding to the production areas were recorded, and bioclimatic indices were calculated as well. Overall, cis-resveratrol concentration was negatively correlated to Huglin index and August mean temperature, whilst positive correlation coefficients were found when considering the Selianinov index and the rainfall of September.