RUSTIONI Laura1, COLA Gabriele2, VANDERWEIDE Josh3, MURAD Patrick3, FAILLA Osvaldo2, SABBATINI Paolo3

1 DiSTeBA – Università del Salento

2 DISAA – Universita’ Degli Studi di Milano

3 Department of Horticulture, Plant & Soil Sciences Building, Michigan State University

Correspondance to:

Article extracted from Laura Rustioni’s presentation in occasion of Enoforum Web Conference (23-25 February 2021)

Phenolic ripening: a matter of oxidation

Tannins play an important role in wine quality and, depending on their chemistry, they can have positive or negative impact. Cellar technologies based on must and wine phenolic oxidation are widespread (Gómez-Plaza & Cano-López, 2011), however, they are not selective with the potential of causing untargeted oxidations. However, a system that modify the oxidative status of seed tannins, without affecting other grape and wine compounds could be pivotal to improve wine chemistry makeup.

Seeds are a primary source of phenolics in grapes and their contribution to the total phenolic content ranges between 0 (for seedless grapes) to 78 % (Rustioni et al., 2019). Seed browning is a major characteristic of the seed ripening process, and it is related to tannin oxidations (Pourcel et al., 2007). Columella (4-70 A.D.) stated that the process of seed darkening is the best grape ripening index in his treatise: “De Re Rustica” (Rustioni & Failla, 2016) and this ancient knowledge is still used by many winegrowers all over the world to decide the best harvesting date.

In optimal growing conditions seed ripening occurs naturally during berry maturation. However, the process can be altered by challenging climate conditions often due to climate change or more often due to specific oenological targets (e.g., low alcoholic concentrations). In those scenarios, tannins have often a sub-optimal quality characteristic.


Our idea: a freeze-thaw treatment

Phenolic decompartmentalization in plant cells cause an oxidative burst (Mohr & Stein, 1969; Holzwarth, Korhummel, Carle, & Kammerer, 2012; Oszmiański, Wojdyło, & Kolniak, 2009). Cells and, especially, vacuoles (the main phenolic storage) are full of water and damaged by freezing events. Thus, we hypothesized that a freeze treatment, damaging the cell membranes, could release phenolics with a consequent oxidation during cell defrost (thaw treatment). The hypothesis was tested in two different viticultural regions: a cool (Michigan) and a warm (Italy). Different cultivars were utilized, and grapes were collected at different ripening stages: 5 in Michigan (the French-American hybrid Chambourcin (Seyval-Villard 12–417×Chancellor), Cabernet Franc, Cabernet Sauvignon, Merlot, Pinot noir) and 6 in Italy (Cabernet Sauvignon, Sangiovese, Syrah, Croatina, Barbera and Nebbiolo). A total of 1400 seeds were analyzed before and after the freeze-thaw treatment (-20°C overnight followed by 3 hours at room temperature).

Figure 1: the effects of the Freeze-thaw treatment on unripen seeds

The results: similarities and differences between natural and artificial seed ripening

Two methodologies (colorimetric and a spectrophotometric) were used to measure seed color modifications during the natural and the artificial (freeze-thaw treatment) seed phenolic ripening. Both methods highlighted similarities and differences between the two processes associated to specific cultivar responses. For example, particularly interesting appeared the results obtained with Nebbiolo grapes.

When seeds are freeze treated at veraison, the phenolic is not completely the same as the one associated to the natural ripening process. Thus, this treatment should not be considered as a substitute of a good vineyard management and optimized genotype x environment interactions: we propose it just as a further possibility of improvement of the tannins’ quality.

Technology for separation of grape seeds at the beginning of winemaking are available (Canales et al., 2008), therefore coupling this technology with the seed freezing treatment could further improve wine quality, helping producers to facing challenging climate conditions during the ripening period. Furthermore, the freeze-thaw treatment does not rely on the use of chemical input (oenological additives), which can be a concern for winemakers.

In our side, we trust in this project and, thus, we are keeping our research interest on this topic. Further details on our experiments are already available in Rustioni et al., 2018 and VanderWeide et al., 2020.


Canales, R., Llaudy, M. C., Canals, J. M., & Zamora, F. (2008). Infuence of the elimination and addition of seeds on the colour, phenolic composition and astringency of red wine. European Food Research and Technology, 226, 1183–1190.

Gómez-Plaza, E., & Cano-López, M. (2011). A review on micro-oxygenation of red wines: Claims, benefits and the underlying chemistry. Food Chemistry, 125(4), 1131–1140.

Holzwarth, M., Korhummel, S., Carle, R., & Kammerer, D. R. (2012). Evaluation of the effects of different freezing and thawing methods on color, polyphenol and ascorbic acid retention in strawberries (Fragaria × ananassa Duch.). Food Research International, 48, 241–248.

Mohr, W. P., & Stein, M. (1969). Effect of different freeze-thaw regimes on ice formation and ultrastructural changes in tomato fruit parenchyma tissue. Cryobiology, 6(1), 15–31.

Oszmiański, J., Wojdyło, A., & Kolniak, J. (2009). Effect of L-ascorbic acid, sugar, pectin and freeze-thaw treatment on polyphenol content of frozen strawberries. LWT – Food Science and Technology, 42, 581–586.

Pourcel, L., Routaboul, J. M., Cheynier, V., Lepiniec, L., & Debeaujon, I. (2007). Flavonoid oxidation in plants: from biochemical properties to physiological functions. Trends in Plant Science, 12(1), 29–36.

Rustioni, L., & Failla, O. (2016). Grape seed ripening evaluation by ortho-diphenol quantification. Italian Journal of Food Science, 28, 510–516.

Rustioni, L., Cola, G., Maghradze, D., Abashidze, E., Argiriou, A., Aroutiounian, R., … Bacilieri, R. (2019). Description of the Vitis vinifera L. phenotypic variability in enocarpological traits by a Euro-

Asiatic collaborative network among ampelographic collections. Vitis – Journal of Grapevine Research, 58(January), 37–46.

Rustioni, L., Cola, G., VanderWeide, J., Murad, P., Failla, O., & Sabbatini, P. (2018). Utilization of a freeze-thaw treatment to enhance phenolic ripening and tannin oxidation of grape seeds in red (Vitis vinifera L.) cultivars. Food Chemistry, 259, 139–146.

VanderWeide, J., Forte, A., Peterlunger, E., Sivilotti, P., Medina-Meza, I.G., Falchi, R., Rustioni, L., & Sabbatini, P. (2020). Increase in seed tannin extractability and oxidation using a freeze-thaw treatment in cool-climate grown red (Vitis vinifera L.) cultivars. Food Chemistry 308, 125571.