A Reference List of Phenolic Compounds (Including Stilbenes) in Grapevine (Vitis vinifera L.) Roots, Woods, Canes, Stems, and Leaves

Due to their biological activities, both in plants and in humans, there is a great interest in finding natural sources of phenolic compounds or ways to artificially manipulate their levels. During the last decade, a significant amount of these compounds has been reported in the vegetative organs of the vine plant. In the roots, woods, canes, stems, and leaves, at least 183 phenolic compounds have been identified, including 78 stilbenes (23 monomers, 30 dimers, 8 trimers, 16 tetramers, and 1 hexamer), 15 hydroxycinnamic acids, 9 hydroxybenzoic acids, 17 flavan-3-ols (of which 9 are proanthocyanidins), 14 anthocyanins, 8 flavanones, 35 flavonols, 2 flavones, and 5 coumarins. There is great variability in the distribution of these chemicals along the vine plant, with leaves and stems/canes having flavonols (83.43% of total phenolic levels) and flavan-3-ols (61.63%) as their main compounds, respectively. In light of the pattern described from the same organs, quercetin-3-O-glucuronide, quercetin-3-O-galactoside, quercetin-3-O-glucoside, and caftaric acid are the main flavonols and hydroxycinnamic acids in the leaves; the most commonly represented flavan-3-ols and flavonols in the stems and canes are catechin, epicatechin, procyanidin B1, and quercetin-3-O-galactoside. The main stilbenes (trans-ε-viniferin, trans-resveratrol, isohopeaphenol/hopeaphenol, vitisin B, and ampelopsins) accumulate primarily in the woods, followed by the roots, the canes, and the stems, whereas the leaves, which are more exposed to environmental stresses, have a low concentration of these compounds. Data provided in this review could be used as (i) a metabolomic tool for screening in targeted and untargeted analyses and (ii) a reference list in studies aimed at finding ways to induce naturally occurring polyphenols on an industrial scale for pant and human disease control.

In V. vinifera, many studies have been published that reported on the concentration of phenolics in berry-containing foods and their impact on human health [18,35]. Indeed, several databases are available for the levels of phenolic compounds in the fruits (e.g., eBASIS, Phenol-Explorer), but none are available for non-edible parts of V. vinifera. Recently, the stems and canes of this economically important genus have been reported as an untapped source of health-promoting compounds [16,17,31,[35][36][37][38][39]. Because of this, numerous efforts for isolation, identification, and quantification of phenolic compounds in the vegetative organs of grapevine have been ongoing. In order to properly design valorization strategies, the precise chemical composition of these vegetative materials has to be known. In this review, a more thorough understanding of the chemical diversity of polyphenols within V. vinifera vegetative organs is provided, which will be useful in this endeavor. The review includes an overview of compounds identified in the roots, cordon and trunk woods, canes, stems, and leaves with their mass and UV spectrum patterns, followed by an estimation of their levels. It concludes with a brief presentation of factors affecting the biosynthesis and accumulation of these compounds. The fallout of such data is multifaceted and will surely contribute to advancing the scientific knowledge in the field.

The Vegetative Organs of the Vine Plant
Grapevine is a climber whose growth in the vineyard is maintained with pruning in order to control the quantity and quality of the grapes [40]. Like any other plant, grapevine has vegetative and reproductive organs. The vegetative organs of vine include the roots and five parts extending from the root system and visible aboveground: trunk, cordons, canes, stems, and leaves. These organs play a key role in light energy capture via photosynthesis, as well as water and nutrient absorption as regulated by transportation.

Roots
The roots of a vine plant are multi-branched structures that grow to various depths into the soil on the basis of the variety (rootstock), and are responsible for anchoring the plant to the ground [12,23,29,30,41,42].

Woods
In the literature, the "wood" refers to samples obtained from the trunk and the cordons. The trunk is composed of sleeves of conductive tissues, most notably the phloem and the xylem [13,[43][44][45]. Cordons or "arms" are extensions of the trunk and the parts where canes (one-year-old wood containing between 8 and 15 buds) and spurs (one-year-old wood containing between two and three buds) originate [12].

Canes
The terms "stems", "canes", "stalks", and "shoots" are sometimes used interchangeably in the literature. For the purpose of this review and on the basis of the literature surveyed, the shoot is the new green growth that develops from buds located on the cordons [24,25,[46][47][48][49]. Once the leaves fall from the vine at the beginning of the dormant season, the brown and harden/woody shoot is considered a cane, which represents a large source of waste derived from the wine industry [40,[50][51][52].

Leaves
Leaves are the most visible parts of the canopy and consist of the blade (the broad, flat part of the leaf designed to absorb sunlight and CO 2 ), and the petiole (the stem-like structure that connects the leaf to the shoot) [4,5,11,19,22,32,[59][60][61][62][63][64].

Extraction, Separation, and Identification of Phenolic Compounds in Grapevine
In grapevine varieties, polyphenols are present as constitutive compounds of the lignified organs (roots, canes, seeds, stems, ripe cluster stems) and/or as induced substances in leaves and berries. In the frame of a long-term project aimed at investigating the physiological and molecular responses of grapevine to trunk diseases [15], several papers that contained the terms "grapevine, grape, vine, vineyard, or vitis" in their titles, plus one of the following terms: "phenolic, polyphenol, flavonoid, anthocyanin, proanthocyanidin, tannin, stilbene, stilbenoid, bioactive, bioactivity, antioxidant, antioxidative, metabolite, metabolic, metabolomic, metabolome, leaf, stem, root, wood, cordon, cane, trunk, phytoalexin, defense, resistance", or terms related to the specific diseases and pathogens of grapevine, were retrieved from citation databases; 80 papers were analyzed that primarily reported on the presence and levels of polyphenols in the vegetative organs (Tables S1 and S2). The term "polyphenol" is used in this review to indicate both the compounds with a second aromatic ring and those arising from the polymerization of flavonoidic/catechin units. Despite their structural diversity, all polyphenols share a common structure element, which consists of a benzene ring to which more than one hydroxyl group is attached [65].
The surveyed literature shows that many extraction methods have been tested, and that several analytical methods using numerous techniques have been developed for the investigation of polyphenols in grapevine, including high-performance liquid chromatography (HPLC) coupled with diode array detection (LC-DAD), HPLC coupled with mass spectrometry (LC-MS, LC-MS/MS), and nuclear magnetic resonance (NMR) [24,28,36,66,67]. Different advantages and disadvantages are associated with each analytical system. Analysis by HPLC-DAD (or HPLC/UV-VIS) is limited by similar or identical absorption maxima of target compounds belonging to the same structural class of polyphenols. Other problems such as lack of baseline resolution, leading to overestimation of individual compound levels, may exist, along with poor sensitivity [16,17,36]. Because of its high selectivity, LC-MS/MS with electrospray ionization (ESI), atmospheric pressure photoionization (APPI), or chemical ionization (APCI) enables the sensitive and simultaneous detection and identification of a large number of (even co-eluting) compounds from a single chromatogram and is therefore the method of choice when libraries are available. MS also enables reductions in the process of sample preparation from extracts [10,68]. NMR, on the other hand, is a non-destructive high throughput method that allows metabolite identification and quantification. It is, however, significantly less sensitive than MS, although more reproducible, especially in long-term studies where samples collected and analyzed over different time periods have to be compared. NMR is also an invaluable tool for the de novo structure determination of compounds [16,59]. In all cases, however, precise conditions are required to achieve a complete qualitative survey of all metabolites over a significant dynamic range in a complex plant extract. Depending on the optimization of extraction and detection parameter settings, two large groups of chemical compounds with phenolic characteristics-that are classified into several structure classes-are clearly delineated in grapevine and are separately discussed in this paper.
The first group comprises phenolic acids (hydroxybenzoic and hydroxycinnamic acids), flavonoids (e.g., flavonols, anthocyanins, proanthocyanidins), and coumarins, which are usually present as preformed compounds in the tissues. Indeed, HPLC in gradient mode on reversed phase C18 columns provides a means to separate most of these compounds in a single chromatography run without the need for derivatization. Due to their structural complexity, however, proanthocyanidins are more easily separated alone by hydrophilic interaction liquid chromatography (HILIC) according their degree of polymerization, or by reverse-phase chromatography, although some of them coelute [69].
The second group is constituted of stilbenic compounds (stilbenes that bear the core structure of 1,2-diphenylethylene and stilbenoids that are hydroxylated derivatives of stilbenes). Several of these compounds are produced naturally by several plants upon attack by pathogens [8,14,70,71]. Because of their dynamic behavior as responses to stresses, the detection of stilbenes requires methods that can be used for monitoring their differential response in various phytopathologic situations [61,71]. Their extraction generally requires specialized instrumentation and expertise, for instance, sample cleaning techniques such as solid phase extraction (SPE), sample dilution, selective extraction, or use of stable isotopes. Most of the qualitative or quantitative analytical studies of stilbenes are performed with HPLC and in an increasing sensitivity order UV, fluorescence (FD), electrochemical (ECD), or MS detection [36,50,52]. A method for the simultaneous separation of proanthocyanidins and stilbenoids has been reported, using a comprehensive bi-dimensional chromatography, with a diol stationary phase in the first dimension and a C18 stationary phase in the second dimension [50,72].

Polyphenols (Excluding Stilbenes) Identified in the Vegetative Organs of Grapevine
Phenolic compounds produced by grapevine range from cell wall-thickening compounds such as lignin and tannins, to specialized compounds such as phenolic acids and flavonoids. The chemical characterization of these compounds is based on analysis of different groups of components individually by LC-MS, mainly in negative ionization mode, although some LC-MS methods in positive-ion mode have been reported [16]. With high-resolution MS, compounds are identified by processing raw data with specific algorithms to calculate molecular formulae on the basis of the monoisotopic mass of the [M-H] − ion and the relative abundances and distances (spacing) of m/z signals measured in the isotopic pattern. Metabolites are then identified by searching in the available MS databases, in comparison with UV spectra patterns reported in the literature [16]. In Table 1, a database specific to grapevine phenolics containing 105 metabolites, including their specific MS and UV information, is provided.

Anthocyanins
Anthocyanins share the same molecular structure of flavonoids composed by one heterocyclic benzopyran ring (as the C ring), one fused aromatic ring (as the A ring), and one phenyl constituent (as the B ring). Nevertheless, they differ on the basis of hydroxyl or methoxyl substitutions in the lateral phenyl B ring, and, in general, for glycosylations and esterifications. Anthocyanins of Vitis are structurally based on five aglycones/anthocyanidins-malvidin, cyanidin, delphinidin, peonidin, and petunidin-which differentiate on the basis of number and position of their hydroxyl groups and their degree of methylation. Acylation occurs at the C6 position of the glucose molecule by esterification with acetic, p-coumaric, and caffeic acids [77,78]. Anthocyanins have been mainly reported in the leaves of grapevine (at least 14) and include: delphinidin (Table 1) [4,5,[77][78][79].

Flavones
Flavones are the simplest members of the class of flavonoids and consist of 4H-chromen-4-one bearing a phenyl substituent at position 2 [65]. Among the flavonoids naturally occurring in grapevine, flavones represent the least common group of aromatic compounds with only apigenin-7-O-glucoside and luteolin-7-O-glucoside reported in the leaves (Table 1) [4].

Flavonols
Chemically, flavonols or 3-hydroxyflavones differ from many other flavonoids in that they have a double bond between positions 2 and 3 and an oxygen (a ketone group) in position 4 of the C ring, like flavones; however, they differ from flavones due to the presence of a hydroxyl group at the position 3. Most of the flavonols exist as O-glycosides and seldomly as C-glycosides, and their conjugated derivatives (glycones) are mainly bound to sugars, hydroxycinnamic acids, or organic acids [35]. Flavonols make up the largest group of flavonoid compounds encountered in grapevine leaves and stems, with at least 35 compounds reported in the literature (Table 1) [1,2,4,5,32,34,59,66,77,79,80] derived from four aglycones: myricetin, quercetin, kaempferol, and isorhamnetin:

Stilbenic Compounds Identified in the Vegetative Organs of Grapevine
The condensation of three C2 residues with an activated hydroxycinnamic acid (as with flavonoids) produces stilbenes, which are metabolites with an essential structural skeleton of two aromatic rings joined by an ethylene bridge (C6-C2-C6) [31,50]. Stilbenes emit a blue fluorescence under UV light with excitation and emission peaks around 320 and 390 nm, respectively [52,63]; in fact, the name "stilbene" derives from the Greek word "stilbos", which is translated as "shining" [16,36]. The chemical structure of stilbenes in both the monomeric and oligomeric states is constituted by a diphenylethylene group oriented in trans or cis. The presence of a cis-stilbenic chromophore gives rise to different spectra, with an absorption maximum of lower intensity and of shorter wavelength compared with that of the trans-isomer [61]. Light exposition of trans-stilbene solutions has been shown to partially photoisomerize stilbenes into cis forms [7,59,61]. There are several areas of confusion with stilbene nomenclature. According to current practice however, the trans/cis nomenclature is used to describe the stereochemistry at saturated rings, whereas the Z/E nomenclature is used to describe the stereochemistry of double bonds [8,13,61,64]. In this review, the trans/cis nomenclature is used, although at least two compounds have been reported with other nomenclatures, namely, miyabenol C and ε-viniferin. Both trans-E-miyabenol C and trans-Z-miyabenol C are reported in the literature [61]. In the case of ε-viniferin, there are two stereochemical centers, at positions 7a and 8a on the dihydrofuran ring, allowing for four potential stereoisomers: (+)-trans-ε-viniferin, (-)-trans-ε-viniferin), (+)-cis-ε-viniferin, and (-)-cis-ε-viniferin) [8].
Most stilbenes have been identified using NMR and MS, which are the most informative techniques. Using MS, the initial identification of compounds is performed on the exact mass measurement of the monoisotopic ion and isotopic pattern, enabling the molecular formula to be identified with a high-confidence score and low mass error. Exact mass measurements of MS/MS fragments either confirm or deny the putative structure. Because MS/MS cannot distinguish between isomeric compounds, tentative assignment is also based on comparisons with data found in the literature [13,16] and NMR profiles. In addition, UV−VIS data based on λmax and UV spectrum when available are compared with those in the literature [12,61]. With this approach, a total of 78 stilbenes have been successfully identified in the vegetative tissues of the vine plant. A database of these stilbenes is provided in Table 2; the masses, when available, are those derived from the negative ion LC−MS datasets. Mass data are usually in agreement among publications, with minor changes in product ions owing to different fragmentation conditions.    4 The most abundant fragments are highlighted in bold. 5 MS = mass spectrometry detection, NMR = nuclear magnetic resonance detection, DAD = diode array or ultraviolet detection. 6 In blue with √ are detected compounds; in light red are undetected compounds or unavailable information.
The basic simple structure of stilbenes gives rise to a wide array of compounds that primarily vary in the number and position of hydroxyl groups and various substitutions with sugars, methyl, and methoxy groups, in addition to the structural conformations of the molecules and oligomerization patterns [8,31,39,68].

Pentameric Stilbenes
Two stilbenes pentamers have been reported in the Vitis genus [39]. However, none have been detected in the vegetative organs.

Levels of Phenolic Compounds in the Leaves, Stems, Canes, Woods, and Roots of the Vine Plant
Several phenolic compounds have been quantified in grapevine, although absolute quantification is currently not available for many of them [9,55,59]. Most authors have reported their data on either a fresh or dry weigh basis (Tables S1 and S2). In analyzing the data for this review, mean values were first calculated using fresh and dry weight values separately, and then together. Although the separate analysis proved challenging because of few data points available for many of the compounds, the final ranking of phenolics was not affected by the method of calculation.
On another note, many of the compounds are quantified as equivalents of the most similar chemicals [12,13,51]. Comparison of the calibration curves showed that assays of content determination in vine of stilbenes in which equivalent chemicals are used as standards lead to a severe underestimation of the oligomer concentration. For example, the quantification of ε-viniferin using trans-resveratrol as standard underestimated its concentration by a factor > 2 in the study by [24]. Therefore, caution is required when interpreting the data presented in Tables 3-10, taking into consideration the water status of the samples analyzed as reported by the different authors, and the number of studies and data points used in the calculation of mean values (Tables S1 and S2).

Phenolic Compounds in Grapevine Leaves
A total of 132 phenolic compounds have been reported in grapevine leaves (Tables 3 and 4 Table 3). These levels in general agree with reports that grapevine leaves are rich sources of flavonols. In their studies, [2,20,22,79,84,85] found that the predominant phenolics in the leaves are quercetin-3-O-glucuronide, quercetin-3-O-glucoside, caftaric acid, and kaempferol-3-O-glucoside; total amounts of quercetin derivatives were significantly higher than total amounts of kaempferol derivatives in the studies [2,79,85]. The 40 stilbenes reported in the leaves of V. vinifera are eminently less abundant than the phenolic acids and flavonoids. The most predominant stilbenes in the leaves with levels superior to 50 mg/kg are trans-resveratrol (137.88 mg/kg), vaticanol C-like isomer (96.85 mg/kg), cis-piceid (78.38 mg/kg), trans-piceatannol (78.04 mg/kg), ampelopsin H (76.08 mg/kg), α-viniferin (71.61 mg/kg), and cis-miyabenol C (50.37 mg/kg) ( Table 4). All these compounds are often undetected in healthy leaves. For example, in two grapevine varieties grown in Serbia, the total stilbene content was 45% higher in infected than in healthy leaf extracts [73]. Interestingly, some leaf samples have been found to contain cis-resveratrol-3-O-glucoside instead of the expected trans-resveratrol as their predominant stilbene [59,60,84].

Distribution of Phenolic Compounds in the Vegetative Organs of Grapevine
A mapping of the distribution of phenolic compounds in grapevine shows that composition and levels greatly vary according to the organ investigated (Figures 1 and 2). In general, flavonoids constitute the largest group of phenolics. From an anatomical point of view, these compounds localize specifically in the stems, with lower amounts in the leaves and the canes (Figure 1).
The overall data obtained through adding together the available individual compound means (not shown) indicate that the total amount of phenolics in the stems is on average 114,415.68 mg/kg. Flavan-3-ols constitute the most abundant group of compounds in the stems (61.63%; catechin, epicatechin, and procyanidin B1 with the highest levels), followed by flavonols (23.75%; quercetin-3-O-galactoside with the highest level), hydroxybenzoic acids (9.03% with a high abundance of gallic acid), hydroxycinnamic acids (3.66%), anthocyanins (0.99%), and stilbenes (0.94%). From an industrial point of view, stems seem to be an important source of proanthocyanidins for potential use as nutraceutical, enological products, chemical standards, or even in winemaking to regulate the composition of flavonoids in wine [69].
Flavonols are quantitatively the most abundant phenolic class in the leaves (83.43% of the total amount of phenolics, i.e., 37,052.70 mg/kg) with a high abundance of quercetin-3-O-glucuronide, quercetin-3-O-galactoside, and quercetin-3-O-glucoside; flavonols are followed by hydroxycinnamic acids (13.19%, of which caftaric acid and coutaric acid have the highest levels), stilbenes (2.63%), flavan-3-ols (0.41%), and hydroxybenzoic acids (0.26%). Coumarins, flavones, anthocyanins, and flavanones are found in minor amounts. The spatial distribution of phenolic compounds in grapevine leaves evaluated by matrix-assisted laser desorption/ionization (MALDI) showed the specific colocation of trans-resveratrol, pterostilbene, and viniferins around the veins in healthy leaves [68]. It is reported that the leaf blade is more abundant in phenolic compounds than the petiole, and much less than the pedicel [75,84].
There are very few reports on flavonols, flavones, flavanones, anthocyanins, and coumarins in the canes. Phenolic groups identified (on average 14,477.42 mg/kg) can be classified in terms of abundance as stilbenes (69.00% of the total) > flavan-3-ols (18.56%; catechin, procyanidin B1 and epicatechin with the highest levels as with the stems) > hydroxycinnamic acids (9.63% of which sinapic acid and ferulic acid) > hydroxybenzoic acids (2.80% of which gallic acid). Indeed, in the comparison of phenolics in the skins, pulps, seeds, canes, and leaves of six cultivars grown in Iran, it was found that the canes usually contained the highest amounts of flavonoids and stilbenes [87].  The data also show that stilbenes accumulate primarily in the woods (34,390.90 mg/kg; ε-viniferin, isohopeaphenol, trans-resveratrol, α-viniferin, and ampelopsin H with the highest levels), followed by the roots (18436.44 mg/kg; ε-viniferin, hopeaphenol, vitisin B, and ampelopsin A with the highest levels), the canes (9989.50 mg/kg; ε-viniferin, trans-resveratrol, isohopeaphenol, vitisin B, and trans-piceatannol with the highest levels), and the stems (1075.55 mg/kg; ε-viniferin, trans-resveratrol, vitisin B, and ampelopsin D with the highest levels), whereas leaves, which are more exposed to environmental stresses [23,40], have a low concentration of these compounds (972.97 mg/kg; trans-resveratrol, vaticanol C-like isomer, piceid, trans-piceatannol, and ampelopsin H with the highest levels) (Figure 2).
Stilbenes are more constitutively expressed in the woods, roots, canes, and stems, where they are believed to help in the prevention of wood rot [12,39,62]. This constitutive expression might explain the more consistent and greater levels of compounds in these plant parts, in contrast to the leaves in which stilbene production is mostly induced to provide enhanced protection against pests and diseases [12,23,65,85]. Levels of stilbenes were compared in the wood, roots, and canes by [12]; grapevine canes usually had ε-viniferin and trans-resveratrol as their main compounds, and woods had more ε-viniferin and isohopeaphenol, whereas roots were generally rich in vitisin B, ampelopsin A, and vitisin A [12]. The authors concluded that the degree of oligomerization of stilbenes increases from the aerial organs to the root system. After manual dissection of the cortex, pith, and conducting tissues of grape canes, the evaluation of the spatial distribution of stilbenes suggested a predominance of monomers in conducting tissues and oligomers in cortex and pith [52].

Factors Affecting the Biosynthesis and Levels of Phenolic Compounds in the Vegetative Organs of Grapevine
Grapevine phenolics occur in large concentration ranges, as attested by standard deviation values presented in Tables 3-10. There are a number of factors that interact together, so as to result in such wide range of phenolic variations. Some of these factors are well documented in the literature and include cultivars, climate, cultural practices, and biotic and abiotic stresses.

Grapevine Cultivars and Rootstocks
Grapevine cultivars are not genetically homogeneous, and most of them are multiplied by vegetative propagation. A collection of vines propagated from the same mother vine make up a clone; clonal selection is routinely carried out in viticulture with the purpose of creating disease-free or high-yielding populations. Progressively, criteria such as the levels of grape sugar and skin phenolic compounds have been integrated in clonal selection programs. Moreover, these clones are often grafted on different rootstocks [2,9,90]. These factors could explain why the contents of polyphenols are subject to such severe variations [6,32,58,59,89]. For instance, plants of Pinot blanc grafted on three different rootstocks-Kober 5B, S04, and 1103P-accumulated resveratrol differentially in the leaves [14]. According to some authors, the best resveratrol-producing cultivars are Pinot noir and Cabernet Sauvignon, depending on the clones investigated [5,6,51]. A comparison of the phenolic profile of canes of the cultivars Chardonnay, Cabernet Sauvignon, Shiraz (Syrah), Merlot, Sauvignon blanc, and Pinot noir showed that Pinot noir had very high levels of trans-resveratrol and trans-ε-viniferin [51]. In several studies, quantitative analyses showed that the stems and leaves of red cultivars are richer in proanthocyanidins, flavonols, hydroxycinnamic acids, anthocyanins, and stilbenes than those of white cultivars [33,53,54,91]. The biosynthesis of phenolic compounds is also closely dependent on plant developmental stages. It has indeed been reported that very young and very old leaves do not synthesize high stilbene and anthocyanin levels, probably because of the incomplete development of the stomata [9,69,77,79,84]. In the study by [89], however, the intensity of stilbene induction did not show a clear and homogeneous correlation with the position of leaves along the shoot. The total viniferin content was generally highest in the second, third, and fourth leaves for the 21/103 genotype. Moreover, [25] did not find a homogeneous trend of change in stilbenoid levels in the stems during the growth cycle.

Geographical Location and Climate (Shading, Temperature, Irrigation)
The geographical location of the vineyard (especially latitude and elevation) and the seasonal meteorological variability in the area are known to influence the phenolic composition of grapevine organs. Meteorological variability, including light, temperature, and water, represents one of the main environmental factors responsible for phenolic biosynthesis. Clear separation was demonstrated between the phenolic profile of leaves [32,79,86] and stems [40] of grapevine cultivars of different geographical origins. A higher variation of stilbene levels between years as compared to variation between plants of the same year have also been reported [89]. Different light exposures of the vine demonstrated that shading decreases the flavonoid content of the leaves, a result that is consistent with the role these molecules play in protecting tissues from UV light [85,91]. In the study by [59], accumulation of quercetin-3-O-glucoside, kaempferol-3-O-glucoside, and quercetin-3-O-galactoside was the most prominent in full sunlight-exposed leaves compared with half-shaded leaves. Furthermore, the biosynthesis of phenolics is sensitive to diurnal differences in temperature, although with different temporal patterns. Indeed, a decrease of flavonoid biosynthesis has been observed when the temperature is limiting or excessive [92]. Extreme weather conditions with prolonged dry periods as well as heavy rain events can severely influence grapevine physiology [2,86]. Water deficit has been reported to upregulate the expression of genes of the anthocyanin pathway [77] and to increase the levels of most polyphenols in the leaves, in particular cis-resveratrol-3-O-glucoside, kaempferol-3-O-glucoside, and quercetin-3-O-glucoside [60]. Recent results show that, during water stress, the synthesis of anthocyanins is paralleled by an increase of the expression of flavonoid transporters [65]. On the other hand, it is reported that excessive water application could induce a decrease in proanthocyanidin levels [92].

Vinicultural Practices
There are many cultural practices that affect the production and accumulation of phenolic compounds in grapevine organs. However, many of these factors seem to act in a typical bell-shaped manner, where they could improve the final levels of compounds only when present at optimal levels. For instance, pruning greatly influences the levels of stilbenoids in the canes, leaves, and stems of grapevine, but the effect depends on the number of branches removed and the duration of the treatment [40,50]. In canes remaining on the plant 30 days after pruning, only a minor increase of total stilbenoid levels was observed, whereas in canes stored at room temperature after pruning, a twofold increase occurred [40]. In general, practices that increase plant vigor, such as fertilizer application, are reported to negatively influence the biosynthesis of phenolics in grapevine. Less trans-resveratrol was accumulated in the leaves when increasing nitrogen doses were applied to one-year-old potted vines, whereas the opposite was observed with increased potassium doses; when nitrogen and potassium were supplied together, potassium did not balance the negative effect of nitrogen [14]. Iron deficiency stimulated anthocyanin accumulation in grapevine apical leaves [78]. The application of other agrochemicals (e.g., plant hormones and chitosan) with the aim of enhancing vegetative growth and grape quality also affects phenolic biosynthesis in a complex way [18]. Typically, it has been reported that abscisic, auxin, and ethylene application lead to an increase in the levels of flavonoids, whereas the opposite has been observed with the application of gibberellic acid and inhibitors of the ethylene receptor [65]. Indirectly, fertilizer and hormone application could also lead to low levels of polyphenols because they induce the production of especially dense foliage that limits the exposure of some organs to sunlight. Grapevine is susceptible to various pests and diseases usually controlled by chemical and biological treatments that can introduce additional variability in the data. High amounts of stilbenoids were produced in grapevine plants that were mycorrhized with Rhizophagus irregularis, as well as an up-regulation in the leaves of genes involved in the stilbene biosynthesis pathway [70].

Outside and Biotic Stimuli
High variability in phenolic levels in grapevine is best explained by biotic stresses and mechanical injuries. In most studies, it was found that the infection status of the plant influences phenolic profiles much more than other factors [9]. Indeed, following pathogen attacks and insect bites, all the vegetative organs of grapevine undergo modifications in terms of their polyphenol composition and contents. For phenolic acids, flavonoids, and coumarins, the literature is contradictory regarding the relationship between level and disease susceptibility [15]. This might be due to the fact that these compounds are part of the constitutive metabolome in lignified tissues. In the study by [77], the expression of flavonoid pathway genes was detected in both healthy and diseased leaves, confirming that the pathway is active in control conditions [77]. These polyphenols that are present prior to an attempted infection of the plant are known as preformed, and are part of a passive resistance mechanism [14,36]. A mechanism of active resistance is the synthesis, degradation, or metabolism to a different compound in response to attacks by pathogens; de novo synthesized compounds are called phytoalexins [71]. An induction in the synthesis of stilbenic compounds in photosynthetic tissues has been considerately reported in response to the main grapevine pathogens, namely, Botrytis cinerea of grey mould [70], Plasmopara viticola of downy mildew [6,10,12,52,61,62,64,68,70,73,82,89], Erysiphe necator of powdery mildew [11,88], fungi associated with grapevine trunk diseases [3,13,15,[43][44][45]86], Rhizopus stolonifera of berry rot [14], Grapevine leafroll-associated virus 3 of Grapevine leafroll disease [77], Xylella fastidiosa of Pierce's disease [9,55], and Aspergillus carbonarius of sour rot [71]. The increment can be as high as 100-fold, and has a biosynthetic origin attributable to stilbene synthase induction. Interestingly, it was found that downy mildew affects the spatial repartition of stilbenoids in the cane, with an increase in the cortex (a tissue notably involved in protection against mechanical damage and microbial attack) and conducting tissues, and a decrease in the pith [52]. Mechanical stress on freshly pruned canes and leaves have also been reported to overinduce the biosynthesis of trans-resveratrol and trans-piceatannol within a short period after pruning [23,40].

Concluding Remarks
Several compounds with phenolic characteristic have been detected in the leaves, stems, canes, woods, and roots of the grapevine plant. An effort is still needed to identify and quantify several of these compounds. It is clear that several factors affect the biosynthetic pathways, leading to the accumulation of phenolic compounds in grapevine. The patterns of gene expression show significant differences between organs and cultivars, especially for genes involved in stilbene synthesis. In the leaves in particular, random inductions in the synthesis of these compounds have been observed, which is understandable given their higher exposure to the environment and resulting susceptibility to attack by pests and diseases. An understanding of the different roles of these factors is crucial because only with this information will it be possible to develop cultural practices aimed at improving phenolic levels in the plants and in the derived products. Moreover, unexplored areas of research related to this topic will most certainly constitute a basis for future improvement of grapevine disease tolerance.