Month: September 2018

Synergistic effect of mixture of two proline-rich-protein salivary families (acidic and basic) on the interaction with wine flavanols

By Alba M. Ramos-Pineda, Ignacio García-Estévez, Montserrat Dueñas, M. Teresa Escribano-Bailón

Astringency is usually defined as the complex of sensations due to shrinking, drawing or puckering of the mouth epithelium (ASTM, 2004). Although the bases of this mechanism are not well understood yet, it is broadly assumed the ability of food tannins to interact with some salivary constituents, mainly salivary proteins, resulting in the formation of protein-tannin aggregates (de Freitas & Mateus, 2012, Rinaldi, Gambuti, & Moio, 2012), which, in turn, causes a decrease in salivary lubrication (Canon et al., 2013).


Salivary proteins have been classified into six groups attending to their structure and characteristics, namely, basic proline-rich proteins, acidic proline-rich proteins, glycosylated proline-rich proteins, statherin, histatins and cystatins (Castagnola, Cabras, Vitali, Sanna, & Messana, 2011; Soares et al., 2011). Among salivary proteins, proline-rich proteins are particularly effective in complexing tannins (Canon et al., 2015). More than 11 human basic proline-rich proteins and five acidic proline-rich proteins isoforms have been identified and the total proline-rich proteins represent >60% by weigh of the total salivary proteome (Inzitari et al., 2005; Messana et al., 2004). The main function proposed for the glycosylated proline-rich proteins is to act as lubricants, whereas proline-rich proteins are associated with tannin precipitation (Oppenheim et al., 2007). In contrast to proline-rich proteins, there are minor structural differences among the acidic proline-rich proteins and their predominant role proposed is related to mineral homeostasis and tooth integrity preservation (Oppenheim et al., 2007).
As aforementioned, the interaction between phenolic compounds and salivary proteins (mainly proline-rich proteins) has been described as key process to explain the astringency perception, with or without precipitation of the tannin-protein complexes (Cala et al., 2012; Ferrer-Gallego et al., 2017). However, studies carried out up till the present have not yet explored the different ability of salivary proteins fractions to interact with flavanols when they are isolated or mixed with other salivary proteins fractions, which could help for unraveling the astringency process.
In this context, this research aimed to study the effect of the coexistence of basic proline-rich proteins and acidic proline-rich proteins salivary fractions on the interaction with flavanols towards the interaction between the same flavanols with these fractions assayed individually, using techniques such as HPLC-DAD, DLS and MALDI-TOF.
To investigate the existence of interactions phenolic compound-salivary protein, determination of the HPLC-DAD profile of salivary proteins before and after the treatment with phenolic compounds (catechin and epicatechin) has been used. Results showed noticeable enhancement of the interaction between (epi)catechin and proline-rich proteins when both types of proteins are blended (see Fig.1), which might indicate that the presence of acidic proline-rich proteins fraction could stabilize the complex formed between proline-rich proteins and flavanols.

Figure 1. Total chromatographic salivary profile (A) showing the different salivary proteins fractions and chromatographic profile of the proline-rich proteins fraction alone (B) or in the sample corresponding to the mixture proline-rich proteins + acidic proline-rich proteins (C), before (blue) and after (red) the interaction with EC. 1: proline-rich proteins; 2: His-3; 3: glycosylated proline-rich proteins; 4: His-3; 5: acidic proline-rich proteins, 6: Cystatins, 7: Statherin and P-B peptide.

To deepen in the characteristics of the possible soluble aggregates formed, the apparent size of the aggregates was determined by dynamic light scattering (DLS) and the identification of the aggregates formed was performed by MALDI-TOF analyses. Results obtained with the different techniques showed the different behaviour of the salivary proteins fractions when they are isolated compared to when blends of salivary proteins are assayed. Up to 30 soluble aggregates were tentatively identified with molecular weight from 4680 to 35,851. (epi)Catechins seem to bind preferentially proline-rich proteins than acidic proline-rich proteins, although the medium size aggregates flavanol- proline-rich proteins formed could favour the interaction with acidic proline-rich proteins giving rise to soluble mixed aggregates (Ramos-Pineda et al., 2019).
Those interested in a longer length report can download the working paper at:


Research Group: (Left to right) Alba M. Ramos Pineda, Dr Ignacio García Estévez, Dr Cristina Alcalde Eon, Dr. Montserrat Dueñas Patón, Rebeca Ferreras Charro, Dr Elvira Manjón Pérez, Dr. M. Teresa Escribano BailónFood Quality Research Team. Grupo de Investigación en Polifenoles (GIP) - Department of Analytical Chemistry, Nutrition and Food Sciences - University of Salamanca (USAL), Salamanca, Spain. Our research projects are focused on the study of the role of phenolic compounds on quality and stability of food products. The main research topic is related to the impact of the phenolic maturity of red grapes on sensory quality of red wines, namely on color and astringency properties. Recent works aimed to go deepen on the study of the mechanism for astringency development through the use of physical-chemical studies, in order to provide the wine industry (both wineries and oenological industries) with basic knowledge about this complex sensation. Furthermore, our studies are also related to the analysis and characterization of phenolic composition (mainly flavonoids and phenolic acids) of different food and plant matrices. This work was carried out in collaboration with Dr. Susana Soares and Dr. Victor de Freitas from the research group of Applied Organic Chemistry (QUINOA) from the Green Chemistry Laboratory (LAQV/REQUIMTE), Department of Chemistry and Biochemistry, University of Porto, Porto, Portugal.





ASTM. (2004). Standard Definitions of Terms Relating to Sensory Evaluation of Materials and Products. (American Society for Testing and Materials, Ed.). Philadephia.
Cala, O., Dufourc, E. J., Fouquet, E., Manigand, C., Laguerre, M., & Pianet, I. (2012). The colloidal state of tannins impacts the nature of their interaction with proteins: the case of salivary proline-rich protein/procyanidins binding. Langmuir, 28, 17410-17418.
Canon, F., Paté, F., Cheynier, V., Sarni-Manchado, P., Giuliani, A., Pérez, J., Durand, D., & Cabane, B. (2013). Aggregation of the salivary proline-rich protein IB5 in the presence of the tannin EgCG. Langmuir, 29, 1926–1937.
Canon, F.; Ployon, S.; Mazauric, J. P.; Sarni-Manchado, P.; Réfrégiers, M.; Giuliani, A.; Cheynier, V. Binding site of different tannins on a human salivary proline-rich protein evidenced by dissociative photoionization tandem mass spectrometry. Tetrahedron 2015, 71, 3039–3044.
Castagnola, M., Cabras, T., Vitali, A., Sanna, M. T., & Messana, I. (2011). Biotechnological implications of the salivary proteome. Trends in Biotechnology, 29, 409–418.
de Freitas, V., & Mateus, N. (2012). Protein/Polyphenol Interactions: Past and Present Contributions. Mechanisms of Astringency Perception. Current Organic Chemistry, 16, 724–746.
Ferrer-Gallego, R., Hernández-Hierro, J. M., Brás, N. F., Vale, N., Gomes, P., Mateus, N., de Freitas, V., Heredia, F. J., & Escribano-Bailón, M. T. (2017). Interaction between Wine Phenolic Acids and Salivary Proteins by Saturation-Transfer Difference Nuclear Magnetic Resonance Spectroscopy (STD-NMR) and Molecular Dynamics Simulations. Journal of Agricultural and Food Chemistry, 65, 6434–6441.
Inzitari, R., Cabras, T., Onnis, G., Olmi, C., Mastinu, A., Sanna, M. T., Pellegrini, M. G., Castagnola, M., & Messana, I. (2005). Different isoforms and post-translational modifications of human salivary acidic proline-rich proteins. Proteomics, 5, 805–815.
Messana, I., Cabras, T., Inzitari, R., Lupi, A., Zuppi, C., Olmi, C., Fada, M. B., Cordaro, M., Giardina, B., & Castagnola, M. (2004). Characterization of the human salivary basic proline-rich protein complex by a proteomic approach. Journal of Proteome Research, 3, 792–800.
Oppenheim, F. G., Salih, E., Siqueira, W. L., Zhang, W., & Helmerhorst, E. J. (2007). Salivary proteome and its genetic polymorphisms. Annals of the New York Academy of Sciences, 1098, 22-50.
Ramos-Pineda, A. M., García-Estévez, I., Soares, S., de Freitas, V., Dueñas, M., Escribano-Bailón, M.T. (2019). Synergistic effect of mixture of two proline-rich-protein salivary families (aPRP and bPRP) on the interaction with wine flavanols. Food Chemistry, 212, 210-215.
Rinaldi, A., Gambuti, A., & Moio, L. (2012). Precipitation of Salivary Proteins After the Interaction with Wine: The Effect of Ethanol, pH, Fructose, and Mannoproteins. Journal of Food Science, 77, 485–490.
Soares, S., Vitorino, R., Osório, H., Fernandes, A., Venâncio, A., Mateus, N., Amado, F., De Freitas, V. (2011). Reactivity of human salivary proteins families toward food polyphenols. Journal of Agricultural and Food Chemistry, 59, 5535–5547.

Posted by in Chemistry, Enology

Inline to online: phenolics measurements made easy

By Jose Luis Aleixandre-Tudo

A research team at the Department of Viticulture and Oenology, Stellenbosch University (DVO) is one step closer to simplification of grape phenolic measurements. The importance of phenolic compounds is due to their crucial role played in the colour and mouthfeel properties of red wines. In a feasibility study recently published, the phenolic content of grapes on a moving conveyer belt was measured using a contactless NIR (Near Infrared) spectroscopy instrument. This inline-online approach was compared against static measurements using the same instrument. Intact as well as crushed berries were measured following the reasoning that phenolic content would be better quantified if grapes are crushed (e.g. the phenolic information of the seed phenolics will be better exposed to the NIR light). The results were compared against two chemical reference methods. The spectral measurements of the crushed berries and the homogenate extraction protocol provided the most accurate prediction models for the inline-online system going forward.

The use of NIR spectroscopy to measure grape phenolics is not new and various studies have reported on it. However, this is the first time where a contactless (no sample preparation) system was tested on a moving conveyer belt in real winery conditions, with very promising results. Such a system can be applied to red grape grading in addition to basic chemical parameters and sanitary status. It can also serve as a decision making tool as to which winemaking practices are needed during the fermentation phase, i.e. amount of pump overs per day, when to press off the skins, enzyme dosage, whether or not to use fermentation tannins, etc. The technology can also potentially be transferred to vineyards to attempt the monitoring of phenolic ripeness during the ripening season.
This project falls into a broader strategy of the research being conducted at the DVO in which efforts are currently being made towards the development of integrated systems to optimise the winemaking process. With the use of spectroscopy (light) applications a number of tools are in the process of being developed, covering from phenolic ripeness and content in grapes to fermentation and ageing monitoring in wines. An example of this can be seen in the South African Phenolics Laboratory (PhenoLAB) web based application that will be launched in 2019. Making use of ultraviolet-visible light and optimised prediction models this application will provide phenolic information in grapes, fermenting samples and wines with minimal analytical time. Other capabilities of the application include the possibility of comparing the generated data against an extensive database of phenolic levels, which can be very valuable for wine producers.

Those interested in a longer length report can download the working paper at:

Aleixandre-Tudo, J.L., Nieuwoudt, H., du Toit, W. Towards on-line monitoring of phenolic content in red wine grapes: A feasibility study. Food Chemistry 270 (2019) 322–331.




Jose Luis Aleixandre-Tudo is currently a researcher in the Department of Viticulture and Oenology at Stellenbosch University (South Africa). After obtaining his PhD in the Universitat Politecnica de Valencia in Spain he moved to South Africa to further investigate the effect of winemaking practices on the phenolic content and evolution during winemaking and aging. His main research focus has shifted towards the development of spectroscopy and chemometrics applications that are later incorporated in a broader process monitoring, control and optimization strategy.


Posted by in Viticulture

Norisoprenoids and aroma precursors in early-harvested grapes

By Maurizio Petrozziello, Andriani Asproudi and Alessandra Ferrandino

The constant increase of summer month temperatures over the last decade has led to significant changes in the chemical composition of grapes at harvest. Many authors have reported a greater and more rapid accumulation of sugars in grapes, and an advance of the phenological phases of flowering and veraison that implies a lack of synchrony among technological, phenolic and aromatic maturity. Considering this evidence early harvesting has gained renewed interest, particularly in the warmer areas of global viticulture. The changes in grape chemical composition that occur close to harvest are crucial for the final quality of the wine and many aspects of the aromatic personality of the grapes are defined in this stage of maturation.

Carotenoids play a crucial role in plants contributing to the photosynthetic process by ensuring the complete uptake of light radiation and protecting chlorophyll from photo-oxidation. Their concentration in grapes depends on several factors namely variety, ripening stage, environmental conditions, including the seasonal meteorological trend, cluster exposure to light, water availability and cultural practices. Some factors facilitate carotenoid synthesis in the herbaceous phase of the berry, while others favour their degradation after veraison and during the final stages of maturation. From an oenological point of view, carotenoids are important precursors of some aromatic compounds known as norisoprenoids that, in turn, play an important role, particularly in wines produced with non-aromatic grapes. Thus, the identification of suitable cultural techniques that may favour both carotenoid synthesis and degradation, as well as the choice of harvest time, appear to be crucial for the quality of future wine.
Norisoprenoids (specifically 13 carbon skeleton compounds) are important contributors to wine aroma due to their very low olfactory thresholds (in water at the ppt level) and pleasant floral and fruity notes, even when present in small quantities. Recent studies show that they can also indirectly contribute to the wine bouquet, acting as enhancers of fruity notes, especially in the case of young wines. Two norisoprenoids are considered key aromas in wine: -ionone and -damascenone. The former is generated directly from both enzymatic and photo-oxidative degradation of β-carotene and contributes a violet-like note to neutral wines. It accumulates in grapes until harvest but its concentration in wines tends to decrease rapidly. The latter confers soft and pleasant notes of rose and quince, and derives from the degradation of neoxanthine via enzymatic and acid-catalysed transformations. The complex pathway and numerous precursors generated explain why -damascenone is generally present in trace amounts in ripe grapes, but it can reach significant concentrations in musts and young wines during their storage.

During a research study carried out in 2015, Barbera and Pinot Noir berries were harvested 7 and 15 days before standard harvest. In contrast to previous research, this study highlighted that during the last stages of ripening, as the berry carotenoid content decreased, a corresponding increase in β-damascenone in wines was not noticed. Moreover, for both Pinot Noir and Barbera, a positive correlation between the total acidity levels of musts and β-damascenone content was highlighted, and the concentration of β-damascenone was higher in wines obtained from early-harvested grapes. In Barbera, a reduction of the alcohol content in the finished wines (grapes harvested 7 days before standard harvest) by approximately 2 degrees corresponded to an increase of β-damascenone concentration in wines of approximately 15%. Thus, for certain varieties, harvesting grapes slightly earlier can have a positive impact on the overall aroma of the produced wine, as early harvesting leads to a reduced alcohol content and is associated with the greater presence of an important aromatic molecule, which is able to enhance the fruity aroma of wines without any effect on the anthocyanin concentration and profile, and thus on wine colour. 

Studying the varietal aromatic profile of a cultivar during the last stages of maturation allows the detection of the optimum aromatic potential of grapes and to understand the relationships among the various classes of compounds, finalizing the choice of time of harvest based on the desired wine type. In light of the above results, it is clear that certain events linking the development of the berry, and the aromatic and technological maturity to the aroma of the final wines are yet to be elucidated. Therefore, the objective of relating grape composition to the aromatic profile of finished wines, especially in the context of climate change, remains a difficult question to approach and highlights the need for further knowledge in this field.

Maurizio Petrozziello

Post-graduate degree in Viticulture and Enology and PhD in Agriculture, Forestry and Food Sciences. Currently he is working as researcher at the Research Centre for Viticulture and Enology of Asti (CREA-VE, Italy). Specialist in GC-MS and GC-O techniques aimed to the analysis of wine odorants. His main interest is aromatic composition of wines and grapes with special attention to winemaking impact on the wine bouquet.


Andriani Asproudi

Graduate degree in Agriculture (Aristotle University of Thessaloniki –Greece), post-graduate degree in Viticulture and Enology and PhD in Agriculture, Forestry and Food Sciences (University of Turin –Italy). Currently she is working as research assistant at the Research Centre for Viticulture and Enology of Asti (CREA-VE, Italy). Her main areas of research are method development and HPLC, GC-MS, GC-O analysis aimed to the characterization of the phenolic, carotenoid and aromatic composition of grapes and wines. Recent research works are focused on the study of the impact of vineyard microclimate and climate change on the evolution of aroma precursors of the grapes and wine key aromas.


Alessandra Ferrandino

(ORCID ID: orcid/org0000-0002-1567-5874)
Born in Turin, 18/10/1968, maiden, two sons.

Her research activity in viticulture and grapevine biology focused on the influences exerted by environment, genotype and their interaction on secondary metabolite accumulation (polyphenols and volatiles). Abiotic and biotic stressors induce grapevine responses able to modify plant hormonal metabolism with consequences on vine morphology, physiology and, finally, concentrations and profiles of secondary metabolites involved in the definition of berry quality. These aspects are the base for understanding the relations existing among grapevine/soil/climate that are responsible for many important practical aspects such as viticulture landscape, wine quality, nutraceutical benefits of grape and derived-product consumption. Moreover, secondary metabolite accumulation in berries and vegetative organs can contribute to explain grapevine susceptibility to pathogens, in the frame of offering new insights to the concept of sustainability in viticulture.

In 2012 she obtained the qualification as Associate professor (ASN, bando 2012, DD n. 222/2012; Sector, 07/B2, Scienze e Tecnologie dei sistemi arborei e forestali; SSD AGR/03, Arboricoltura generale e coltivazioni arboree – Fruit tree cultivation).
She is author/co-author of more than 100 technical/scientific papers; 33 are indexed on Scopus and ISI Web of Science (Scopus: h index = 13; 781 total citations).
She has developed scientific collaborations with IPSP - CNR (Turin, Italy); CREA (Asti, Italy); Istituto Fisica Applicata Nello Carrara - CNR (Firenze, Italy); Institut Agricole Régional (IAR, Aosta, Italy); Technical University of Munich (Biotechnology of Natural Products and Fruit Science, Freising, Germany); University of Castilla-La Mancha, Catédra de Química Agricola Albacete, Spain; ICVV Instituto Ciencia Vid y Vino, Llogrono, Spain.
She is reviewer of international Journals such as Journal Agriculture and Food Chemistry, Food Chemistry, Food Research International, Sensors, HortScience, OenoOne, Phytochemistry, to cite a few.
She held educational activities: Fruit and Grapevine cropping systems - Orchard management for fruit quality (Viticulture); Master in Agricultural Science (academic years 2013-14; 2014-15; 2015-16; 2016-17; 2017-18).
Secondary metabolites in grapevine. Inter-university Master in Viticulture and Enology Sciences (academic years 2014-15; 2015-16; 2016-17; 2017-18).
Interdisciplinary activities; Degree in Viticulture and Enology (2015-16).
PAS and TFA courses for teacher qualification in 2013-14 and 2014-15, respectively.
She was supervisor/co-supervisor of Master Degree Thesis and co-tutor of PhD students at DISAFA, Turin University. She took part in activities related to Incoming Student Guidance; she is deputed to manage Incoming and Outcoming Students from and to Bordeaux and Montpellier.


Those interested in a longer length report can download the working paper at:


Asproudi A., Petrozziello M., Cavalletto S., Guidoni S. (2016). Grape aroma precursors in cv. Nebbiolo as affected by vine microclimate. Food Chemistry, 211:947-956.
Baumes, R., Wirth, J., Bureau, S., Gunata, Y., & Razungles, A. (2002). Biogeneration of C 13-norisoprenoid compounds: experiments supportive for an apo-carotenoid pathway in grapevines. Analytica Chimica Acta, 458(1), 3-14.
Bindon K.A., Dry P.R., Loveys B.R. (2007). Influence of plant water status on the production of C13-norisoprenoid precursors in Vitis vinifera L. cv. Cabernet Sauvignon grape berries. J Agric Food Chem, 55: 4493-4500.
Lee S.H., Seo M.J., Cotta M.J.P., Block D.E., Dokoozlian N.K., Ebeler S.E., (2007). Vine Microclimate and Norisoprenoid Concentration in Cabernet Sauvignon Grapes and Wines. Am. J. Enol. Vitic., 58, 291-301
Longo, R., Blackman, J. W., Antalick, G., Torley, P. J., Rogiers, S. Y., & Schmidtke, L. M. (2017). Harvesting and blending options for lower alcohol wines: a sensory and chemical investigation. Journal of the Science of Food and Agriculture.
Oliveira C., Csar Ferreira A., Costa P., Guerra J., Guedes de Pinho P., 2004. Effect of Some Viticultural Parameters on the Grape Carotenoid Profile. J Agric Food Chem, 52 (13): 4178-4184.
Palomo, E. S., Díaz-Maroto, M. C., Viñas, M. G., Soriano-Pérez, A., & Pérez-Coello, M. S. (2007). Aroma profile of wines from Albillo and Muscat grape varieties at different stages of ripening. Food control, 18(5), 398-403.
Pineau, B., Barbe, J.-C., Van Leeuwen, C., & Dubourdieu, D. (2007). Which Impact for β-Damascenone on Red Wines Aroma? Journal of Agricultural and Food Chemistry, 55(10), 4103–4108.
Young, P. R., Lashbrooke, J. G., Alexandersson, E., Jacobson, D., Moser, C., Velasco, R., & Vivier, M. A. (2012). The genes and enzymes of the carotenoid metabolic pathway in Vitis vinifera L. BMC Genomics, 13(1), 243.

Posted by in Enology, Viticulture

Scale effect of viticultural zoning: effect of macro-terroir and basic terroir unit in Chianti Classico D.O.C.G. (Italy)

By Simone Priori

The terroir effect on wine is one of the most debated issue in the world of wine, because of the complicated interaction between natural and human factors. Beside the human factor, which plays the most important role through viticultural and oenological practices, the soil and climate factors are most important in the expression of terroir, and they may vary depending on the spatial scale. At regional scale, climate in interaction with the grapevine cultivar is the most important (Jones et al., 2005), whereas at “wine district scale”, the interaction between mesoclimate, topography and geology might be the dominant factor for grape and wine peculiarities (Van Leeuwen et al., 2004; Priori et al., 2014; Ramos et al., 2015). At farm and vineyard scale, the micro-climate, the soil features, especially the soil hydrology, become much more important to differentiate the grape peculiarities (Costantini et al., 2009; Bramley et al., 2011; Priori et al., 2013; Bonfante et al., 2015; Tardaguila et al., 2017). The object of this post is to present the results of a very recent paper aimed to investigate the effect of macro-terroir and basic terroir units (UTB) on wine peculiarities under three contrasting vintages (Priori et al., 2019).

The vineyards studied for this work belong to Barone Ricasoli estate, one of the largest and oldest farms in the Chianti Classico wine district, Tuscany. These vineyards are situated in three of the most important and representative macro-terroir (MT) of the Chianti Classico wine districts (Fig. 1), which are: i) CALC- calcareous, clayey and stony soils on hills at medium altitude (350-450 m a.s.l.) characterized by calcareous flysch (limestone and shales) of the Monte Morello Fm.; ii) SAND- sandy and stony soils, non-calcareous, on hills at medium altitude (400-450 m a.s.l.), characterized by sandstone of Macigno fm. (SAND); iii) MAR- sandy-loamy soils, at lower altitude (300-350 m a.s.l.) formed from marine sandy gravelly deposits of the Pliocene period. In addition, a macro-terroir more specific for this farm was selected on ancient fluvial terraces (FLUV) characterized by calcareous, loamy and stony soils at low altitude (250-300 m a.s.l.).


Figure 1. The four macro-terroir (MT) studied for this work, with the main soil features.

The study was carried out in 12-16 years old Sangiovese cv. vineyards, the vine density varies between 6200 and 6600 vines/ha, and the trellis system was the simple spurred cordon with vertical shoot. The experimental vineyards received the same viticultural treatments during the growing season. Each MT was subdivided into two 1.5-2 ha size UTB by cluster analysis, using proximal soil sensing data obtained by electromagnetic induction sensor and gamma-ray spectrometer (Fig.2). Apparent electrical conductivity (ECa), measured by electromagnetic induction sensor, is more related to soil texture, stoniness, and soil depth until depth of about 1.5 m, whereas gamma-ray spectra are related to parent material mineralogy, soil texture, stoniness, and calcium carbonate of the topsoil (0-30 cm). Therefore, the couples of UTB within each MT were mainly distinguished by texture, stoniness, and then associated hydrological features like available water capacity (AWC) and internal drainage.
Within each UTB, three representative soil profiles of about 1.5 m were dug, to characterize the main soil features and grapevine root distribution. Small areas with deep and too fertile soils, or subject to waterlogging or exceptional soil erosion, were excluded. The grape from each UTB was harvested and vinified separately, using the same methods, in a tank of 80 hl and then in 5 hl oak barrel (tonneaux) for a 6-months aging. The wines were analysed and evaluated by a panel of 10 wine tasters through a “blind tasting”. Sensory analysis was mainly comparative, aimed to assess the differences among the wines. The wine tasters gave a score, ranging from 1 to 10 to several wine parameters, giving score 10 to the wine which best expressed the parameter. In addition, the tasters indicated their feeling about the wine in terms of aroma typology (fruity, floral, spicy, herbaceous), using 0-absent, 1-scarce, 2-medium, and 3-strong.



Figure 2. Workflow of the UTB mapping for selective harvest.

The study was carried out in 12-16 years old Sangiovese cv. vineyards, the vine density varies between 6200 and 6600 vines/ha, and the trellis system was the simple spurred cordon with vertical shoot. The experimental vineyards received the same viticultural treatments during the growing season. Each MT was subdivided into two 1.5-2 ha size UTB by cluster analysis, using proximal soil sensing data obtained by electromagnetic induction sensor and gamma-ray spectrometer (Fig.2). Apparent electrical conductivity (ECa), measured by electromagnetic induction sensor, is more related to soil texture, stoniness, and soil depth until depth of about 1.5 m, whereas gamma-ray spectra are related to parent material mineralogy, soil texture, stoniness, and calcium carbonate of the topsoil (0-30 cm). Therefore, the couples of UTB within each MT were mainly distinguished by texture, stoniness, and then associated hydrological features like available water capacity (AWC) and internal drainage.
Within each UTB, three representative soil profiles of about 1.5 m were dug, to characterize the main soil features and grapevine root distribution. Small areas with deep and too fertile soils, or subject to waterlogging or exceptional soil erosion, were excluded. The grape from each UTB was harvested and vinified separately, using the same methods, in a tank of 80 hl and then in 5 hl oak barrel (tonneaux) for a 6-months aging. The wines were analysed and evaluated by a panel of 10 wine tasters through a “blind tasting”. Sensory analysis was mainly comparative, aimed to assess the differences among the wines. The wine tasters gave a score, ranging from 1 to 10 to several wine parameters, giving score 10 to the wine which best expressed the parameter. In addition, the tasters indicated their feeling about the wine in terms of aroma typology (fruity, floral, spicy, herbaceous), using 0-absent, 1-scarce, 2-medium, and 3-strong.
The climatic conditions were very variable during the three experimental vintages, from very warm and dry (2012), on average with long term climate (2013), and colder and more humid than the average (2014). The contrasting climate conditions that occurred during the three experimental years determined vintage-to-vintage differences in the grape and wine quality (Fig.3). The coldest and most humid summer of 2014 provided poor differentiations between wines, hiding the terroir effect, at MT scale and, especially between the couples of UTB. On the contrary, during the warmest and driest summer of 2012, the wines from the UTB couple within each MT showed the clearest differences.
In general, macro-terroir tends to have stronger effect on must pH, wine total acidity, glycerine and colour intensity, whereas climate conditions of the vintage stronger influence must malic acid, wine total polyphenols, anthocyanins and dry extract (Tab.1). A large amount of variance was also explained by the interaction between MT and climate of the vintage, especially dry extract and colour intensity.



The effect of UTB sub-division of the studied vineyards showed influence in wine colour and flavour intensity. Changing the zoning scale from MT to UTB according to soil physical and hydrological properties, appeared to have a significant effect only under dry summers, like in 2012 and, to a less extent, in 2013.
From the oenological and wine tasting results, the wine produced in the different UTB, belong to the four MT, showed the following peculiarities:

  • Both the UTB situated in soils developed on clay-calcareous flysch (CALC) provided wines with general higher alcohol, total anthocyanins, dry extract and colour intensity than the average. Previous studies reported the same results for wines produced in the same macro-terroir (Scalabrelli et al., 2001; Priori et al., 2013).
  • Loamy-sandy soils, non-calcareous, developed on feldspathic sandstone (SAND), provided wines characterized by light colour intensity and low acidity. SAND2, characterized by higher stoniness, and then soil macroporosity and faster drainage, provided wines with higher fruity flavour and general higher flavour intensity.
  • Loamy-sandy soils, calcareous, developed on marine deposits (MAR) provided wines with low (MAR1) or medium (MAR2) alcohol, as well as low glycerine and dry extract. Moderate water deficit, which characterized MAR2 during the vintages 2012 and 2013, seems to play a key role to increase the flavour intensity of the wine, in particular for fruity and floral notes.
  • Loamy and clay-loamy soils, calcareous, developed on ancient fluvial terraces (FLUV) made wines richer in alcohol, polyphenols, and glycerine, as well as high colour intensity. The effects of the two UTB within this MT is not still clear and strongly variable according to the vintage climate.



Table 1. Fischer’s F-values resulting from the mixed-design ANOVA, using MT effect as fixed factor, vintage effect as random factor, and UTB nested in MT. In bold, F-values significant for p < 0.05.

Figure 3. The main must and wine parameters in the 8 UTB during the three vintages. The lines represent the average value of that vintage.

In conclusion, geology, soilscape and mesoclimate features, which characterize MT, drive some major wine peculiarities over time, while soil physical-hydrological features, which typify UTB within the same MT, play a key role on wine distinctiveness, mainly during dry vintages. The use of a more robust delineation of homogeneous areas, carried out using innovative techniques like proximal soil sensors, is fundamental to discriminate UTB. On the other hand, management of scattered and irregular UTB areas for selective harvest can be very difficult, or almost impossible, without a precision viticulture approach involving mechanical harvesters and GNSS.

I want to thank all the colleagues who collaborated to this work, in particular: Edoardo Costantini, Sergio Pellegrini, and Giuseppe Valboa (CREA-AA, Agriculture and Environment, Florence) and Paolo Storchi, Rita Perria, and Sergio Puccioni (CREA-VE, Viticulture and Eonology, Arezzo). Moreover, special thanks to “Barone Ricasoli” s.p.a. farm, who supported the work through VignaCRU project, and to the agricultural manager of the farm Massimiliano Biagi and co-workers (Fabio Cascella, Marco Cerqua, and Claudio Carapelli) for their support.
Those interested in a longer length report can download the working paper at:



Simone Priori ( is a researcher at CREA-AA, Research Centre of Agriculture and Environment in Florence (Italy), in the research group of “Database and Digital Soil Mapping Laboratory” ( He received his Ph.D. from the University of Siena, Department of Earth Science in 2009, and since 2012 have permanent position at CREA. His research focused on pedology, digital soil mapping, and proximal soil sensing, especially applied to viticultural zoning and precision viticulture. He was vice-coordinator and project manager of the international project Core-Organic+ “Resolve” and scientific leader of several research projects funded by private stakeholders (wineries, consultants). He has published more than 25 papers in international scientific journals, many of which on the role of the soil in grape quality, soil management in vineyard, digital soil mapping for precision viticulture.




Bonfante, A., Agrillo, A., Albrizio, R., Basile, A., Buonomo, R., De Mascellis, R., ... & Manna, P., 2015. Functional homogeneous zones (fHZs) in viticultural zoning procedure: an Italian case study on Aglianico vine. Soil, 1(1), 427.
Bramley, R. G. V., Ouzman, J., Thornton, C., 2011. Selective harvesting is a feasible and profitable strategy even when grape and wine production is geared towards large fermentation volumes. Australian Journal of Grape and Wine Research, 17(3), 298-305.
Costantini, E.A.C., Pellegrini, S., Bucelli, P., Storchi, P., Vignozzi, N., Barbetti, R., Campagnolo, S., 2009. Influence of hydropedology on viticulture and oenology of Sangiovese vine in the Chianti area (Central Italy), Hydrol. Earth Syst. Sci. Discuss., 6, 1197-1231.
Jones, G.V., White, M.A., Cooper, O.R., Storchmann, K., 2005. Climate change and global wine quality. Climatic Change, 73,3, 319-343.
Priori, S., Martini, E., Andrenelli, M.C., Magini, S., Agnelli, A.E., Bucelli, P., Biagi, M., Pellegrini, S., Costantini, E.A.C., 2013. Improving wine quality through harvest zoning and combined use of remote and soil proximal sensing. Soil Sci. Soc. Am. J., 77(4), 1338-1348.
Priori, S., Barbetti, R., L’Abate, G., Bucelli, P., Storchi, P., Costantini, E.A.C., 2014. Natural terroir units, Siena Province, Tuscany. Journal of Maps, 10-3, 466-477.
Priori, S., Pellegrini, S., Perria, R., Puccioni, S., Storchi, P., Valboa, G., Costantini, E. A., 2019. Scale effect of terroir under three contrasting vintages in the Chianti Classico area (Tuscany, Italy). Geoderma, 334, 99-112.
Ramos, M.C., Jones, G.V., Yuste, J., 2015. Phenology and grape ripening characteristics of cv Tempranillo within the Ribera del Duero designation of origin (Spain): Influence of soil and plot characteristics. European Journal of Agronomy, 70, 57-70.
Scalabrelli, G., D’Onofrio, C., Ducci, E., Bertuccioli, M., 2001. Grapevine performances in five areas of Chianti Classico. Rev.S.Vitic. Arboric. Hortic. 33: 253-260.
Tardaguila, J., Diago, M.P., Priori, S., Oliveira, M., 2017. Mapping and managing vineyard homogeneous zones through proximal geoelectrical sensing. Archives of Agronomy and Soil Science, 64(3), 409-418.
Van Leeuwen, C., Friant, P., Chone, X., Tregoat, O., Koundouras, S., Dubourdieu, D., 2004. Influence of climate, soil, and cultivar on terroir. American Journal of Enology and Viticulture, 55(3), 207-217.

Posted by in Viticulture

Understanding the green character in red wines by a sensory-directed approach

By Sara Ferrero del Teso, María-Pilar Sáenz Navajas, Ignacio Arias Pérez, Purificación Fernández Zurbano, Ana Escudero, Vicente Ferreira

Flavour in food and beverages is the result of sensory interactions between sensory active volatile and non-volatile molecules (Prescott, 2012). An increase, in knowledge about flavour formation would allow to have objective tools to manage the production process and optimise the quality of the final product. The chromatographic separation of volatile and non-volatile compounds or group of compounds followed by sensory description of fractions is a common strategy that flavourists use to identify sensory-active molecules related to both aroma and taste or mouth-feel properties (Sáenz-Navajas et al., 2017; Scharbert, Holzmann, & Hofmann, 2004).
Nevertheless, the study of the sensory activity of non-volatile molecules is less explored than volatiles. The absence of reference materials with defined mouthfeel properties, makes it difficult to progress in finding the chemical compound or group of compounds responsible for such properties in complex foods or beverages such as wine. Furthermore, the study of the sensory activity of non-volatile molecules, takes more time and resource consuming especially due to sensory fatiguing induced by in-mouth tasting in comparison with orthonasal olfaction needed to evaluate aroma-related molecules.

The presented work has applied the flavourist strategy of separating both volatile and non-volatile molecules, to focus on fractions driving green character in red wines. It is known, that during last years, due to climate change, there is a difference in time between technological (related to sugar and acidity content) and phenolic maturity of grapes. Based on declarations of winemakers, the fact that technological maturation is achieved, while phenolics and aroma precursors are still unripe, lead to green wines, which is associated with a decrease in consumers’ acceptance of the product (Sáenz-Navajas et al. 2017). Experts, recurrently use the term green to describe certain wines, but the term “green character” seems to be an ill-defined term. This project firstly aimed at understanding what experts mean when they use “green” to describe a flavour.

In this context, our work, focused on firstly defining the ill-defined term: green character, and then isolating and identifying the group of compounds (volatiles and non-volatiles) inducing such property. Finally, the real sensory implication, of these compounds or group of compounds, on the green character, was confirmed by spiking real wine samples. Thirty-eight wines with a priori different levels of green character, based on winemakers’ suggestions, were submitted to descriptive analysis by a panel of fifteen wine experts. Twelve descriptive attributes were rated on a five-point scale: six related to taste and mouthfeel properties (green tannin, dry tannin, astringency, sourness, sweetness and soft tannin) and six aroma-related (intensity, woody, vegetal, fresh fruit, ripe fruit and oxidation/reduction). Besides, two multidimensional terms, preference and green character, were also rated. Results showed that green character was a multidimensional term negatively correlated to preference and to woody aroma, sweetness and soft tannins, while positively to vegetal aroma, astringency, green and dry tannins (Figure 1).


Figure 1. PCA with descriptors as active variables and green character and preference as supplementary variables

Based on these results, for the study of sensory activity of non-volatile molecules, two wines with the highest scores in green character (W1 and W2) and other two with the lowest (W3 and W4) were selected. The non-volatile fraction of these four samples was fractionated by semipreparative liquid chromatography followed by solid-phase extraction. Six non-volatile fractions (Figure 2) were isolated for each wine and submitted to sensory characterization (together with the four selected wines) focused on taste and mouthfeel properties. Sixteen winemakers were provided with a list of 23 terms, which was previously generated in an intensive sensory work developed in a previous step. It consisted in a Rate-all-that-Apply (RATA) strategy as described in Sáenz-Navajas et al. 2017. That is, they had firstly to find the terms from the list that described the samples and only for those, rate the intensity in a seven-point-scale.


Figure 2. Scheme of wine fractionation

Based on the results of the descriptive analysis, for the study of sensory activity of volatile molecules, one wine with high (W5) and one (W6) with low vegetal aroma and green character were selected. The aroma extracts of both wines were fractionated by preparative liquid chromatography. The aroma properties of the 15 fractions obtained were classified by orthonasal evaluation employing a sorting task. However, no fraction with green-related aromas was found. Thus, another set of 16 wines were scored for green character. Quantitative characterization of sensory-active volatiles revealed that green wines were especially higher in isoamyl alcohol.
The sensory impact of the two non-volatile fractions (F13 and F22+F23) and volatile isoamyl alcohol on the green character was evaluated in two very different wines: a young (YW) and an oaked (OW) red wine. The wines, originally displaying no green character, were spiked with F22+F23, F13, isoamyl alcohol and submitted to sensory analysis by a panel of fifteen wine experts (Figure 3).



Figure 3. Sensory confirmation analysis

Interestingly, the single addition of isoamyl alcohol, proanthocyanidins (F22+F23) or anthocyanin-derivative compounds (F13) to a young or oaked aged red wine (base samples) does not generate a significant increase of the green character. However, the presence of isoamyl together with one of the two phenolic fractions significantly increase green character of wines. These results confirm that the interaction between isoamyl alcohol and the anthocyanin-derivative fraction and/or tannins is involved in the formation of green character in red wines by increasing astringent-related properties of wines. However, we were not able to find any volatile compound increasing the vegetal aroma of green wines. This result is very interesting firstly, because it shows that the volatile compound isoamyl alcohol, which has a spirit-like aroma can modulate mouthfeel properties and also because it confirms the mouthfeel properties elicited by anthocyanins, which was already observed in independent studies of our research group in Sáenz-Navajas et al. (2017) and of other groups by surface plasmon resonance (Guerreiro et al., 2017). Besides, results showed that young wines present significantly higher scores than oaked wines for green character. This result confirms that the green character is wine dependent and suggests that the oaked, ripe fruit and oxidation aromas of the oaked wine could mask green character.
Those interested in a longer length report can download the working paper at:



Institute of Grapevine and Wine Sciences (ICVV).
Oenology Research Group: Chemical and sensory analyses in enological research, Logroño, Spain.
The group studies the role of the aroma and the flavor of the wines. The research is carried out in collaboration with the Laboratory for Aroma Analysis and Enology (LAAE) from the University of Zaragoza. Understanding flavor and mouthfeel sensations are our main focus. We addressed this research by combining strategies of separation (chromatography), technologies of chemical elucidation (mass spectrometry) and organoleptic analysis (sensory analysis).
Research Group: Purificación Fernández Zurbano, María-Pilar Sáenz-Navajas, Marta Dizy Soto, José Federico Echávarri Granado, Sara Ferrero del Teso, Shu Yan Liu.




Ares, G., Bruzzone, F., Vidal, L., Cadena, R. S., Giménez, A., Pineau, B., Jaeger, S. R. (2014). Evaluation of a rating-based variant of check-all-that-apply questions: Rate-all-that-apply (RATA). Food Quality and Preference, 36, 87-95.
De-La-Fuente-Blanco, A., Sáenz-Navajas, M. P., & Ferreira, V. (2016). On the effects of higher alcohols on red wine aroma. Food Chemistry, 210, 107-114.
Ferreira, V., Hernández-Orte, P., Escudero, A., López, R., & Cacho, J. (1999). Semipreparative reversed-phase liquid chromatographic fractionation of aroma extracts from wine and other alcoholic beverages. Journal of Chromatography A, 864(1), 77-88.
Gonzalo-Diago, A., Dizy, M., & Fernandez-Zurbano, P. (2013). Taste and Mouthfeel Properties of Red Wines Proanthocyanidins and Their Relation to the Chemical Composition. Journal of Agricultural and Food Chemistry, 61(37), 8861-8870.
Gonzalo-Diago, A., Dizy, M., & Fernández-Zurbano, P. (2014). Contribution of low molecular weight phenols to bitter taste and mouthfeel properties in red wines. Food Chemistry, 154, 187-198.
Guerreiro, J.R.L., Teixeira, N., De Freitas, V., Sales, M.G.F. , & Sutherland, D.S. A saliva molecular imprinted localized surface plasmon resonance biosensor for wine astringency estimation. Food Chemistry, 233, 457-466.
Harbertson, J., Picciotto, E., & Adams, D. (2003). Measurement of polymeric pigments in grape berry extracts and wines using a protein precipitation assay combined with bisulfite bleaching. American Journal of Enology and Viticulture, 54(4), 301-306.
Revelette, M. R., Barak, J. A., & Kennedy, J. A. (2014). High-performance liquid chromatography determination of red wine tannin stickiness. Journal of Agricultural and Food Chemistry, 62(28), 6626-6631.
Sáenz-Navajas, M. P., Avizcuri, J. M., Ferrero-del-Teso, S., Valentin, D., Ferreira, V., & Fernández-Zurbano, P. (2017). Chemo-sensory characterization of fractions driving different mouthfeel properties in red wines. Food Research International, 94, 54-64.
Sáenz-Navajas, M.P., Arias, I., Ferrero-del-Teso, S., Fernández-Zurbano, P., Escudero, A., Ferreira, V. (2018). Chemo-Sensory Approach for the Identification of Chemical Compounds Driving Green Character in Red Wines. Food Research International 109, 138–148.

Posted by in Chemistry, Enology, Food Science and Technology