Introduction

Wine composition reflects the applied environmental, viticultural and oenological practices. Thus, wines from diverse viticultural areas, using selected yeasts in alcoholic fermentation or even berries from different clones, can result in wines with distinct sensorial characteristics. Such variability, due to monoterpene compounds (Duchêne et al., 2009), thiol precursors (Nicolini et al., 2019) and other classes of varietal aroma compounds, is of great interest to winemakers. Studies on vine clones aim to evaluate the adaptation, production and also search for differentiated aromatic characteristics. The viability of a clone in a given location is strongly influenced by the environment (soil, climatic conditions and vineyard management techniques) and depends on the final use of the produced grapes and regulatory circumstances (Keller, 2015). For this reason, studies related to the competition of clones in new growing areas are essential.

In recent years, clones of the aromatic cultivars Gewürztraminer, Moscato Giallo and Müller Thurgau have been studied to evaluate the adaptation to specific regions and their contribution to free compounds and precursors in berries and wines (Nicolini et al., 2013; Nicolini et al., 2016; Nicolini et al., 2019).

Chardonnay, a variety from Burgundy, has 31 certified clones in France (Plantgrape). Some of them are aromatic (clones 809, 1068, 1145, and 1146; Plantgrape) due to the volume of monoterpenoid compounds biosynthesised through MEP (methyl-erythritol-phosphate) and MVA (mevalonic acid) pathways. In a genetic study of the grape and wine aroma (Lin et al., 2019) of an aromatic clone of the Chardonnay cultivar (cv.), the authors observed an increase in monoterpenoid synthesis due to a mutant gene that expresses the enzyme 1-deoxy-D-xylose-5-synthase (DXS). Such an alteration suggests a potential catalysation for substrate build-up to produce monoterpenes through the MEP pathway.

In addition to the terpenoid pathways noted by several authors (Slaghenaufi et al., 2020; Yue et al., 2020), starting with isoprenoids units (C5) and the enzymes that contribute to the biosynthesis of these compounds in plants, Rienth et al. (2021) described how the biosynthesis of such aromatic compounds varies depending on berry stage. Peak production occurs during the herbaceous development phase (Phase I) of the fruit, which decreases until véraison (Phase II) and increases again during ripening (Phase III). Papers have described how these compounds are accumulated during the developmental stages in a cool Australian climate region in the following cultivars: Shiraz, Cabernet-Sauvignon, Riesling, Chardonnay and Pinot gris (Zhang et al., 2016; Luo et al., 2019). However, no data have yet been reported for Brazil´s tropical and subtropical regions.

Since Chardonnay clone 809 has more distinctive aromatic characteristics, which have already been evaluated by Duchêne et al. (2009) in France, this work aims to investigate the use of this material in the south of Minas Gerais (Brazil) to produce a Chardonnay sparkling wine with distinct aromatic traits and to compare the volatile compounds formed in all the process steps to Chardonnay clone 76, a non-aromatic material.

Materials and methods

Results

3. Multivariate analysis

For the multivariate analysis, 84 and 90 compounds in the base wines and 89 and 93 volatiles in the sparkling wines were identified in 2017 and 2018 vintages respectively.

Figure 1. Principal component analysis (PCA) for volatile compounds identified in the base (BASE W) and sparkling wines (SPARK) of Chardonnay clones (CL) 76 and 809 in two consecutive vintages in southeast Brazil (Caldas/MG).

Save View full size Expand inline Collapse inline

Figure 1A: PC1 vs PC2, clusters differentiating the process step (left: base wine and right: sparkling wine) and the clones (bottom: clone 76 and top: clone 809). Figure 1B: Four clusters in PC3 – left and right sides the base wines of 2017 and 2018, respectively, and in the middle top: the sparkling wines of clone 809, and bottom: sparkling wines of clone 76.

In total, 121 variables were noted and considered significant using Tukey´s test (p < 0.05). It was thus possible to carry out the PCA, in which the ratio of the areas of the free compounds (compound area/internal standard area) was determined for clones 76 and 809. The total variance of the data for the three principal components was 74.2 % (Figure 1A and 1B). PC1 differentiated the elaboration stage by 42.3 % (base vs sparkling wine), and PC2 discriminated the clones used to prepare the base and sparkling wines (clone 809 vs 76; PC2: 16.8 %). For the third component, there were four clusters; the base wines were differentiated by the vintage (2017 base wine vs 2018 base wine). However, the sparkling wines were clustered by their clones (Figure 1B). The most significant volatile compounds in the sparkling wines of clone 809 that differentiated them in both vintages were α-terpineol, linalool, ß-myrcene, hotrienol, nerol oxide and limonene.

Discussion

This is the first study to be conducted on the characterisation of volatile composition of Chardonnay clone 809 compared to clone 76 in Serra da Mantiqueira, southeast Brazil (Caldas, MG), a region where Chardonnay clone 76 is generally used in commercial vineyards A mutation in the enzyme 1-deoxy-D-xylose-5-synthase catalyses the production of monoterpenoid compounds in the Chardonnay clone 809, which may impart a more floral and fruity character to the wine.

Differences in the volatile composition of the grapes were found between harvests; this can be explained by the variability of the climatic conditions, which depend on the season, temperature, rainfall, thermal amplitude and solar radiation, and which affect enzymatic activity, gene expression and primary and secondary metabolism (Drappier et al., 2017; Luo et al., 2019; van Leeuwen et al., 2020).

Regarding the volatile composition in the other steps, statistical differences were found between the main compounds (esters, volatile fatty acids and higher alcohols) in the base wine between clone and season, which can be explained by the lack of esterification (volatile fatty acids with ethanol or higher alcohol with acetyl-CoA) in 2017 due to poor enzymatic activity, ethanol amount or ester hydrolysis due to temperature. However, the volume of esters in the sparkling wines for both clones was not found to be statistically different, demosntrating that an equilibrium was reached by the main volatile compound class in the final product.

The percentage of TDA of monoterpenoid compounds in clone 809 (aromatic clone) grapes is higher (Table 1), on the other hand the results for clone 76 corroborate data reported in the literature (Duchêne et al., 2009; Luo et al., 2019); this demonstrates that Chardonnay is a cultivar that contains a low quantity of these compounds, with most of its clones being considered non-aromatic. The scents produced by the main monoterpenoid compounds found in the berries in both seasons (e.g., linalool, α-terpineol, citronellol, nerol and geraniol) are floral, sweet and citric. An evaluation could be carried out on the viticultural practices that favour the synthesis of terpenoid compounds, such as exposing bunches to light (Friedel et al., 2016; van Leeuwen et al., 2020), and which consequently enhance the wine bouquet.

The difference in free monoterpenoid compounds in the musts comparing the harvests for clone 809 may be due to the influence of environmental conditions (light and temperature) on enzymatic activity and gene expression, as previously described regarding the volatile composition of berries.

The reduction in terpenoid content from the must to base wine (13.3 % TDA decrease) in both seasons can be explained by the base wine's tartaric and protein stabilisation stages. Using bentonite for protein clarification can reduce aromatic compounds in sparkling wine production (Ubeda et al., 2021). In addition to eliminating proteins, bentonite removes aromatic compounds; it is not selective when used as a clarifier to eliminate proteins or as an adjuvant in the remuage process. Additionally, Slaghenaufi et al. (2020) found that these compounds can be adsorbed by yeast cells due to their polarity and thus be lost. Further studies should be conducted on how to avoid the loss of terpenoid compounds during sparkling wine production (fining, adjuvant and adsorption in dead yeast cell walls).

In 2018, monoterpene esters were evaluated in the base wine of both clones. Although yeasts from the Saccharomyces genus are unable to biosynthesise terpene compounds due to the lack of terpene synthase enzymes (González-Barreiro et al., 2015), monoterpenol compounds can undergo biotransformation catalysis through acetylation, forming terpinyl acetates (citronellyl acetate from β-citronellol, and geranyl acetate from geraniol), as demonstrated by Slaghenaufi et al. (2020) using the Saccharomyces cerevisiae strain.

Monoterpene alcohols can be converted by reduction, oxidation, hydroxylation, isomerisation, translocation and cyclicisation (King and Dickinson, 2000; Slaghenaufi et al., 2020) into other terpenoid compounds; for example, geraniol can be converted into citronellol or linalool. In this study, the lack of geraniol in the base and sparkling wines may be one explanation for the conversion to other monoterpenoids, but further studies are necessary to confirm this. Although this conversion can influence the organoleptic properties and the intensity of wines (King     and Dickinson, 2000), geraniol (antibacterial and cytotoxic) conversion is a way the yeast enzymes detoxify the media (Slaghenaufi et al., 2020).

Regarding the multivariate analysis, Figure 1B suggests that, regardless of the vintage, the sparkling wines in the third component had an influence of the clones. Thus, even after two fermentations, the rose and sweet character of the sparkling wine produced with clone 809 may be due to the following monoterpenoid compounds: α-terpineol, linalool, ß-myrcene, hotrienol, nerol oxide and limonene. The alcohols of monoterpenes are potent odorants (González-Barreiro et al., 2015) which may confer floral, fruity, sweet and pungent aromas (Burdock, 2010) to the wine.

Conclusion

The data suggest that clone 809 expresses a varietal characteristic due to the monoterpenoid compounds in the berries and sparkling wines, differing from clone 76 in terms of quantities of terpenoid compounds. Some of these compounds discriminated the sparkling wines made with Chardonnay clone 809 in the multivariate analysis, primarily monoterpene alcohols, which are considered fairly odoriferous. Among the monoterpene alcohols, compounds such as linalool, α-terpineol and hotrienol are present, which have flowery, fruity and sweet aromas.

This preliminary study in the Serra da Mantiqueira region could be a driver for further experiments on improving the aromatic potential of clone 809 in the vineyard and during vinification, applying, for example, agricultural techniques (e.g., training systems and exposure of bunches and leaves to solar radiation) and different vinification processes (e.g., use of other yeasts, develop the process in such way that the loss of terpenoid compounds is reduced). Additionally, extraction methods could be applied to evaluate the free and bound forms of the volatile compounds and to quantify them in order to determine the precise potential of the use of this clone for sparkling wines.

Acknowledgements

Laboratory infrastructure: the University of São Paulo and FAPESP (Grant number: 2013/07914-8); Vineyard implementation: FAPEMIG (Grant number: CAG-PPM-00301-16) and Vitacea Bra.

This investigation was supported by laboratory practices (infrastructure, equipment and consumables) and the vineyard implementation in the agronomical centre. Thanks to all EPAMIG staff who helped in the vineyards and winery.

References

• Burdock, G.A. (2010). Fenaroli´s handbook of flavour ingredients. Pp. 2135. Florida, USA: CRC Press. https://doi.org/10.1201/9781439847503
• Carlin, S., Vrhovsek, U., Franceschi, P., Lotti, C., Bontempo, L., Camin, F., Toubiana, D., Zottele, F., Toller, G., Fait, A., & Mattivi, F. (2016). Regional features of northern Italian sparkling wines identified using solid-phase micro-extraction and comprehensive two-dimensional gas chromatography coupled with time-of-flight mass spectrometry. Food Chemistry, 208, 68-80. https://doi.org/10.1016/j.foodchem.2016.03.112
• Drappier, J., Thibon, C., Rabot, A., & Geny-Denix, L. (2017). Relationship between wine composition and temperature: Impact on Bordeaux wine typicity in the context of global warming – Review. Critical Reviews in Food Science and Nutricion, 59(1), 14-30. https://doi.org/10.1080/10408398.2017.1355776
• Duchêne, E., Legras, J. L., Karst, F., Merdinoglu, D., Claudel, P., Jaegli, N., & Pelsy, F. (2009). Variation of linalool and geraniol content within two pairs of aromatic and non-aromatic grapevine clones. Australian Journal of Grape and Wine Research, 15, 120-130. https://doi.org/10.1111/j.1755-0238.2008.00039.x
• Friedel, M., Frotscher, J., Nitsch, M., Hofmann, M., Bogs, J., Stoll, M., & Dietrich, H. (2016). Light promotes expression of monoterpene and flavonol metabolic genes and enhances flavour of wine grape berries (Vitis vinifera L. cv. Riesling). Australian Journal of Grape and Wine Research, 22, 409–421. https://doi.org/10.1111/ajgw.12229
• González-Barreiro, C., Rial-Otero, R., Cancho-Grande, B., & Simal-Gandara, J. (2015). Wine aroma compounds in grapes: A critical review. Critical Reviews in Food Science and Nutrition, 55(2), 202–218. https://doi.org/10.1080/10408398.2011.650336
• Ilc, T., Werck-Reichhart, D., & Navrot, N. (2016). Meta-analysis of the core aroma components of grape and wine aroma. Frontier Plant Science, 7, 1-15. https://doi.org/10.3389/fpls.2016.01472
• Keller, M. (2015). The science of grapevines: anatomy and physiology. Pp. 509. Washington, USA: Elsevier Inc. ISBN: 978-0124199873
• King, A., Dickinson, J.R. (2000). Biotransformation of monoterpene alcohols by Saccaromyces cerevisiae, Torulaspora delbrueckii and Kluyveromyces lactis. Yeast, 16, 499-506. https://doi.org/10.1002/(SICI)1097-0061(200004)16:6<499::AID-YEA548>3.0.CO;2-E
• Lin, J., Massonnet, M., & Cantu, D. (2019). The genetic basis of grape and wine aroma. Horticulture Research, 6, 1-24. https://doi.org/10.1038/s41438-019-0163-1
• Luan, F., Mosandl, A., Gubesch, M., & Wüst, M. (2006). Enantioselective analysis of monoterpenes in different grape varieties during berry ripening using stir bar sorptive extraction- and solid phase extraction-enantioselective-multidimensional gas chromatography-mass spectrometry. Journal of Chromatography A, 1112(1-2), 369–374. https://doi.org/10.1016/j.chroma.2005.12.056
• Luo, J. Q., Brotchie, J., Pang, M., Marriott, P. J., Howella, K., & Zhang, P. Z. (2019). Free terpene evolution during the berry maturation of five Vitis Vinifera L. cultivars. Food Chemistry, 299, Article 125101. https://doi.org/10.1016/j.foodchem.2019.125101
• Nicolini, G., Moser, S., Borin, G., Tonidandel, L., Román, T., & Larcher, R. (2013). Gli aromi del Moscato Giallo nelle sue interpretazioni in Trentino e nei Colli Euganei. Rivista L´enologo, 2, 65-72. ISSN: 0392-176X
• Nicolini, G., Román Villegas, T., Slaghenaufi, D., Moser, S., Malacarne, M., Larcher, R., & Malossini, U. (2016). Müller Thurgau: nuovi cloni italiani di una varietà Mittel-Europea. Rivista Internet di Viticoltura ed Enologia. URL http://www.infowine.com. Accessed 02/01/2021. ISSN 1826-1590
• Nicolini, G., Malossini, U., Tonidandel, L., & Román, T. (2019). Differenze clonali nella dotazione di precursori dei tioli varietali in Gewürztraminer. Rivista Internet di Viticoltura ed Enologia. URL http://www.infowine.com. Accessed 02/01/2021. ISSN 1826-1590
• Plantgrape. Chardonnay. Retrieved from http://plantgrape.plantnet-project.org/en/cepage/Chardonnay. Accessed December 19, 2020.
• Rienth, M., Vigneron, N., Darriet, P., Sweetman, C., Burbidge, C., Bonghi, C., Walker, R.P., Farniani, F., & Castellarin, S.D. (2021). Grape berry secondary metabolites and their modulation by abiotic factors in a climate change scenario – a review. Frontiers in Plant Science, 12, Article 643258. https://doi.org/10.3389/fpls.2021.643258
• Schwab, W., Davidovich-Rikanati, R., & Lewinsohn, E. (2008). Biosynthesis of plant-derived flavor compounds. Plant Journal, 54, 712-732. https://doi.org/10.1111/j.1365-313X.2008.03446.x
• Slaghenaufi, D., Indorato, C., Troiano, E., Luzzini, G., Felis, G.E., & Ugliano, M. (2020). Fate of grape-derived terpenoids in model systems containing active yeast cells. Journal of Agricultural and Food Chemistry, 68(47), 13294-13301. https://doi.org/10.1021/acs.jafc.9b08162
• Sun, Q., Gates, M.J., Lavin, E.H., Acree, T.E., & Sacks, G.L. (2011). Comparison of odor-active compounds in grapes and wines from Vitis vinifera and non-foxy American grape species. Journal of Agricultural and Food Chemistry, 59(19), 10657-10664. https://doi.org/10.1021/jf2026204
• Ubeda, C., Lambert-Royo, M. I., Cortiella, M. G., Barrio-Galán, R., & Peña-Neira, A. (2021). Chemical, physical, and sensory effects of the use of bentonite at different stages of the production of traditional sparkling wines. Foods, 10(2), Article 390. https://doi.org/10.3390/foods10020390
• Ugliano, M., & Henschke, P.A. (2009). Yeast and Wine Flavour. In: Wine Chemistry and Biochemistry (edited by M. V. Moreno-Arribas, & M. C. Polo). Pp. 313 – 392. New York, USA: Springer Science+Business Media. https://doi.org/10.1007/978-0-387-74118-5
• van Leeuwen, C., Barbe, J., Darriet, P., Gerroy, O., Gomès, E., Guillaumie, S., Helwi, P., Laboyrie, J., Lytra, G., Menn, N.L., Marchand, S., Picard, M., Pons, A., Schüttler, A., & Bei, R.D. (2020). Recent advancements in understanding the terroir effect on aromas in grapes and wines. OENO One Vine Wine, 54(4), 985-1006. https://doi.org/10.20870/oeno-one.2020.54.4.3983
• Yue, X., Ren, R., Ma, X., Fang, Y., Zhang, Z., & Ju, Y. (2020). Dynamic changes in monoterpene accumulation and biosynthesis during grape ripening in three Vitis vinifera L. cultivars. Food Research International, 137, Article 109736. https://doi.org/10.1016/j.foodres.2020.109736
• Zhang, P., Fuentes, S., Siebert, T., Krstic, M., Herderich, M., Barlow, E. W. R., & Howell, K. (2016). Terpene evolution during the development of Vitis Vinifera L. cv. Shiraz grapes. Food Chemistry, 204, 463–474. https://doi.org/10.1016/j.foodchem.2016.02.125