This study aimed to investigate flavan-3-ol composition under altered microclimatic conditions in Cabernet Sauvignon grown in the Stellenbosch Wine of Origin District.
Proanthocyanidins or condensed tannins are mostly situated in the solid parts of the cluster (skins, seeds and stems) and to a lesser degree in the pulp (Sun et al., 1999; Jordão et al., 2001; Ó-Marques et al., 2005). Proanthocyanidins are transferred from the solid parts of the grape (skins, seeds and stems) into the must during winemaking operations (crushing, maceration and fermentation). They are partly responsible for the wine organoleptic properties (Monagas et al., 2003). The quantity, structure and degree of polymerisation of grape proanthocyanidins differ, depending on their localisation in the grape tissues (seeds, skin and pulp). Seeds have the highest concentration of procyanidins (Ricardo da Silva et al., 1992a). Within the grape berry, proanthocyanidins or condensed tannins are situated in the hypodermal layers of the skin and the soft parenchyma of the seed between the cuticle and the hard seed coat (Adams, 2006).
Furthermore, a higher degree of polymerisation occurs in grape skins (Adams, 2006). In berry skin, (+)-catechin has been identified as the main terminal and extension subunit skins (Escribano-Bailón et al., 1995; Souquet et al., 1996; De Freitas et al., 2000; Kennedy et al., 2001; Downey et al., 2003; Obreque-Slier et al., 2010). The main flavan-3-ol subunits present in grape seeds are (+)-catechin, (−)-epicatechin and (−)-epicatechin-3-O-gallate. The main terminal subunit is (+)-catechin and the main extension subunit is (−)-epicatechin (Romeyer et al., 1986; Prieur et al., 1994; Downey et al., 2004). Grape skins differ from seeds, because (−)-epigallocatechin and a lower proportion of galloylated units are present in the skins (Escribano-Bailón et al., 1995; Souquet et al., 1996; De Freitas et al., 2000; Kennedy et al., 2001; Downey et al., 2003; Obreque-Slier et al., 2010).
Materials and methods
The site and methods have been discussed in Part 1 of this series. Additionally the compositional analysis of proanthocyanidins was carried out following acid-catalysed cleavage in the presence of excess phloroglucinol (phloroglucinolysis) (Kennedy & Jones, 2001a). The method provided information regarding the subunit composition, mean degree of polymerisation (mDP), percentage of galloylation (%G) and the percentage of prodelphinidin units (%P) in grape skins and seeds where applicable. The proanthocyanidin cleavage products were determined by RP-HPLC using a method adapted from Kennedy and Taylor (2003).
Results and discussion
Seed tannin compositional data by phloroglucinolysis revealed that the terminal seed flavan-3-ol subunits were (+)-catechin, (−)-epicatechin and (−)-epicatechin-3-O-gallate (Table 1). The proportional composition of terminal subunits changed throughout berry development in both seasons (data not shown). The seasonal impact on the seed proanthocyanidin terminal subunit composition was larger than the treatment impact. This is primarily due to the higher light intensities observed in the 2010/2011 season and lower light intensities in the 2011/2012 season.
(−)-Epicatechin was the main constituent of the seed extension subunits with (+)-catechin and (−)-epicatechin-3-O-gallate being present in lower proportions in both seasons (Table 2). The proportional composition of extension subunits changed throughout berry development in both seasons. The results of this study agree with that of Fujita et al. (2007) and Cohen et al. (2008), who reported minimal variation in the seed proanthocyanidin composition with shading, heating and cooling of berries.
Seed mDP varied between 2.7 and 8.8 in the 2010/2011 season and 2.9 and 7.7 in the 2011/2012 season during berry development among treatments (data not shown). In the 2011/2012 season, the respective treatments had higher amounts of extension subunits at the beginning of berry ripening than the 2010/2011 season, resulting in higher mDP (data not shown). The decrease of mDP and avMM from fruit set were observed.
The percentage of galloylated derivatives was determined during both seasons (Table 3). A significantly higher (p ≤ 0.001) percentage of galloylation was observed in the STD treatment when compared with the other three treatments in 2010/2011 (Table 3). During the 2011/2012 season, no significant differences were observed between the galloylation percentages among the treatments, suggesting that galloylation was influenced more by the season than the applied treatments (Table 3).
Grape skin compositional changes during ripening
(+)-Catechin, (−)-epicatechin and (−)-epicatechin-3-O-gallate were identified as the grape skin proanthocyanidin terminal subunits (Table 4). (+)-Catechin was the predominant compositional contributor, with epicatechin and (−)-epicatechin-3-O-gallate present in lower proportions or not detected (Table 4). There was a significant difference (p ≤ 0.001) in the mean (+)-catechin and (−)-epicatechin-3-O-gallate terminal subunit contribution between the two seasons (Table 5).
(−)-Epigallocatechin was the predominant extension subunits followed by epicatechin in grape skins. Lower levels of (+)-catechin and (−)-epicatechin-3-O-gallate were found in both seasons (data not shown). In this study, light exposure did not have a significant impact on the extension unit composition in either of the seasons investigated.
In the 2011/2012 season, the lowest average polymer length was observed in the LR (-UV-B, 2xUHI) treatment, while the LR (-UV-B,-PAR) (shaded without leaves and laterals) treatment showed higher mDPs (Table 6). The low mDP observed in the LR (-UV-B, 2xUHI) treatment can be a result of the high PAR (Blancquaert et al., 2019) that may have been above optimal levels. However, this is not supported by LRW treatment that had similar high PAR values combined with higher temperatures in 2010/2011. Chorti et al. (2010) reported that excessive sunlight exposure could result in excessive sunburn, which could influence skin proanthocyanidins in the grape berry. Where similar mDP values were obtained between the seeds in the respective seasons, skin mDPs were higher in 2011/2012 when compared with 2010/2011 (Table 6).
From this study, it can be concluded that tannin composition is influenced by complex interactions within a particular season. These interactions include seasonal climatic patterns and particularly the light quality and quantity from flowering until harvest which has a direct impact on the berry size and seed number, as well as the accumulation of flavonoids. There was no clear impact of treatment and thus light quality and quantity on either seed tannin content or concentration (Part 1). In the case of skin tannin there was an indication of increased skin tannin with light exposure, but this was only visible in the 2010/2011 seasons indicating that seasonal variability had a larger impact then the individual treatments applied to alter the light quantity and quality. Therefore, seasonal differences should be taken into account.
These results contribute to a better understanding of the light and seasonal interaction on flavonoid composition. Furthermore, additional studies performed in a greenhouse or growth chamber, to control the temperature and light conditions to have a clearer understanding of the abiotic factors influencing flavonoid composition in the berry, could be beneficial.
– For more information, contact Erna Blancquaert at firstname.lastname@example.org.