3.2.2. Extraction during winemaking

recently reported in Merlot grapes and wines [67]. Flavan-3-ols occur in several isomeric forms. The carbons at the C2 and C3 positions of the flavan-3-ol backbone are two asymmetric centers, such as the five monomeric flavan-3-ols are grouped into two diastereomers pairs, with configurations 2R:3S for (+)-catechin and (+)-gallocatechin and 2R:3R for (-)-epicatechin, (-)-epigallocatechin, and ()-epicatechin-3-O-gallate [68]. These different isomeric configurations, in turn, have an impact on bioavailability [69], antioxidant and radical scavenging properties [70], and, ultimately, on sensory properties [71, 72], as further discussed later.

In seeds, flavan-3-ols are located in thin-walled cells between the external cuticle and the inner lignified layers. The (sometime observed) seed browning during berry ripening is thought to be the result of both monomeric flavan-3-ols and tannins undergoing oxidation [73]. Seeds contain only (+)-catechin, ()-epicatechin, and (-)-epicatechin-3-O-gallate [64, 74]. In the skins, flavan-3-ols occur in the subepidermal cell as shapeless or spherical inclusions free in the vacuoles but also associated with the tonoplast [75]. Skins contains (+)-catechin, (-)-epicatechin, (-)-epicatechin-3-O-gallate, and, additionally, (-)-epigallocatechin [76], as well as trace amounts

Quantitative differences also occur within the berry tissues. Seeds concentrate the vast majority of flavan-3-ols of the berry [66, 75–79]. For example, flavan-3-ol concentrations of about 179 mg/kg fresh weight (FW) have been found in Cabernet Sauvignon seeds whereas skins of the same variety only have 4.8 mg/kg FW [80]; similar results are reported for other varieties [81, 82]. In wines, the content of monomeric flavan-3-ols varies from 29 to 41 mg/L in Tempranillo and Graciano wines [78], 107 to 176 mg/L in Pinot Noir wines [83], 182 mg/L in

Due to the reduced state of the C ring of the flavan-3-ol structure, and thus favorable oneelectron donation properties, flavan-3-ols can react with several wine electrophiles. The condensation of monomeric flavan-3-ols with anthocyanins either by a direct covalent reaction between them [85, 86] or mediated by acetaldehyde [87, 88] is one of the main reactions with impact on

of (+)-gallocatechin and (-)-epigallocatechin gallate [66, 77].

Figure 7. Monomeric flavan-3-ols found in V. vinifera grapes and wines.

162 Phenolic Compounds - Natural Sources, Importance and Applications

Tannat wines [84], and up to 288 mg/L in Cabernet Sauvignon wines [43].

color during wine aging and is further addressed in Section 3.4.

Release of flavan-3-ols from skins occurs early during winemaking, within the first 2 or 3 days of skin contact [107–110]. For example, levels of (+)-catechin and (-)-epicatechin after 5 days of skin contact represented between 80 and 85% of the maximum content attained later at pressing in Grenache wines [60]. In Tempranillo and Grenache-Graciano blends, the release of flavan-3-ols occurred early, between days 3 and 5 of maceration, and remained unchanged during a postmaceration period of 1 week followed alcoholic fermentation [109]. In this same study, (-)-epigallocatechin, only located on the skins, was extracted rapidly, suggesting that the extraction of flavan-3-ols and small oligomers from skins occurs during the first days of maceration [109].

Release of flavan-3-ols from seeds requires longer maceration times. For example, the maximum extraction of flavan-3-ols from seeds occurred after 2 to 3 weeks of maceration in model wines, and under those extended conditions, the seeds contributed almost 90% of the total flavan-3-ol content of the final wines [108]. Also, in model wines containing only seeds of the variety Monastrel, the levels of (+)-catechin and (-)-epicatechin increased from 5 mg/L at day 2 to 27 mg/L at day 10 [111]. This represents a slower rate of extraction relative to that of flavan-3-ols from skins. Moreover, longer maceration times, favored by the winemaking technique known as extended maceration, enhance the overall extraction of flavan-3-ols and, more specifically, that of epicatechin-3-O-gallate from seeds [43] (Figure 8b). Other studies have also reported that the percentage contribution of galloylated subunits increases along with maceration time [109, 111–113], and this, together with the content of epigallocatechin (exclusive of the skins), has been used as a surrogate to estimate the percentage contribution of seeds (and skins) to the overall wine's flavan-3-ol and tannin content [114].

Since extraction of flavan-3-ol from seeds occurs toward the end of a regular maceration period (e.g., 15 days), the extraction of both flavan-3-ols and tannins from seeds was assumed to be mediated by the dissolutive effect of ethanol on the lipidic outer coat of the seeds, which will typically occur toward the end of the fermentation process [115]. From this perspective, higher ethanol levels should selectively favor the extraction of seed phenolics in general and that of flavan-3-ols in particular [48, 110]. However, model wine experiments have shown that extraction of both flavan-3-ols and tannins can occur in the absence of ethanol, and that the role of ethanol may be to solely increase the rate of extraction [111]. Current evidence supports the notion of maceration length, and not alcohol, as being one of the main drivers of seed tannin extraction, at least under typical final ethanol concentrations (13–14.5%, v/v) [47, 116]. Thus, an alternative explanation for the late release and extraction into wine of seed flavan-3-ols and tannins is that extraction from the seeds may only occur after the seeds had attained maximal hydration. In studying seed extraction in model wine solutions, and noticing a lag phase

Figure 8. Monomeric flavan-3-ol concentration and composition and tannin size distribution by concentration and composition in Cabernet Sauvignon wines grouped as a function of the maceration length treatment. (a) Control wines (10 days of maceration) and (b) extended maceration wines (30 days of maceration). CE: catechin equivalents. Adapted from [43].

during the time course of maceration, Singleton and Draper noted that "…The lag may be an expression of the fact that most cells do not surrender their constituents for extraction while they are still living. Swelling and osmotic-pressure phenomena may also be involved…" [117]. If this is accepted, then the onset of seed tannin and flavan-3-ol extraction will occur once seeds have reached full hydration, whereby the leakiness of the parenchyma cells outside the true seed coat would allow the prompt release of flavan-3-ols [111, 118].
