Aging of Malbec Wines from Mendoza and California: Evolution of Volatile Composition

The search for strategies to improve the quality of Argentine Malbec wines has led to experimentation with production of wines that achieve superior sensory properties after prolonged bottle aging. During aging, a series of complex oxidation-reduction and polymerization reactions occur. These reactions, responsible for changing the sensory aspects of wines, can be influenced by a variety of factors (terroir, winemaking practices, storage conditions, etc.). Thus, understanding the chemical processes that take place during bottle aging could significantly impact the enol-ogy industry. This work evaluates the evolution of aromatic profiles over time in Malbec wines from Mendoza and California.


Introduction
Bottle aging can improve the sensory properties of some wines (Gambuti et al. 2013).This process can range from a few years to more than a century, depending on the nature and composition of the wine.While some wines evolve favorably over time, many others are made to be consumed young.The property that defines whether a wine can withstand long aging is the aging potential (AP), defined as the ability of a given wine to improve its sensory quality over time (Jaffré et al. 2009).
Argentina is the largest producer of Malbec wines in the world, with 86% of its production in the province of Mendoza.During the 20th century, this variety was established as the insignia of the country.The incessant search for new tools and strategies that allow the local wine industry to continue to improve the quality of Argentinian Malbec has led winemakers to experiment with production of age-worthy wines.
Terroir, understood as the environmental, edaphic, and cultural factors of a grapegrowing region (Baker and Clarke 2012), can have a great impact on the chemical composition and sensory attributes of young wines and, consequently, on their AP.In turn, factors associated with winemaking (pH, antioxidant concentration, access to O 2 , phenolic composition, and ripening conditions, among others) and external factors (aging temperature, closure type, bottle size, humidity, light exposure, etc.) are expected to also have a major impact on sensory changes.
The evolution of a wine over time can be divided into three stages, during which various electron transfer (oxidation-reduction), hydrolysis, condensation, and polymerization reactions take place.These reactions, which change the sensory aspects of wines, are initiated by the catalytic activation of dissolved O 2 .In the presence of tartaric acid and other ligands, and complexes of Fe (II) and possibly other metal cations (Cu, Mn, Cr, Co, Ni), an auto-oxidative process is triggered, leading to formation of various oxidation products (among them acetaldehyde), which become involved in subsequent polymerizations between

Methods and key findings
Wines from Mendoza and California were obtained under standardized conditions.Determination of volatile profiles was carried out on young wines by gas chromatography-mass spectrometry and, subsequently, after seven years of aging.The effect of time and region on volatile profiles was investigated.The aging time showed a significant effect on the aromatic composition.While the profiles obtained after seven years were less diverse than at bottling, they retained a significant number of desirable volatile compounds.Additionally, the volatile profiles of the aged wines still permitted differentiation of samples by region of origin.

Conclusions and significance
The information obtained enabled us to assess the evolution of aromatic profiles during bottle aging of Malbec wines from Mendoza and California, and could be useful in decision-making regarding the blending, marketing, and storage of these wines.flavanols and anthocyanins (Fulcrand et al. 2006).In the first stage, known as development, chemical changes occur that improve wine sensory quality, characterized by reduced astringency and increased color stabilization.During the second maturity stage, the wine reaches its maximum sensory impact, which is the ideal time for consumption.Finally, a period of deterioration begins, in which the wine loses color and body, often becoming drying on the palate (Ribéreau-Gayon et al. 2006, Linsenmeier et al. 2022).
Storing a bottle of wine for an adequate time allows it to reach its full sensory impact: a more rounded palate and the development of aromatic bouquet.Aromas associated with wood, vanilla, dried fruit, chocolate, and cooked fruit (Ribéreau-Gayon et al. 2006) replace the aromas of young wines (fruity and floral), and color stabilizes from intense purple to reddish hues (Ribéreau-Gayon et al. 2006).
Currently, it is not possible to objectively determine the storage time and conditions necessary to achieve maximum sensory improvement from wine composition, therefore, the future AP of a wine is estimated using sensory analysis.A new strategy for estimating AP has been proposed; applying cognitive definitions from professional tasters regarding the aging potential of champagne reserve wines, this strategy is based on a three-dimensional scale and sensory analysis method (Le Menn et al. 2021).Estimating wine resistance to oxidation is also critical to assess wine aging capacity (Waterhouse and Miao 2021).The information gathered from such an approach, while useful, provides enologists with only a partial view of the aging process.Consequently, studying the link between initial composition and the chemical changes that occur during wine aging is considered central to understanding this process and to predicting AP.
Several authors have investigated the evolution of physicochemical indicators with storage time.The aromatic profile as determined by a panel of tasters provided some clues to define the aging capacity of Burgundy wines (Langlois et al. 2010).This study determined that young wines with low acidity and high astringency had greater AP.It concluded that the concept of vin de garde (from French, referring to a wine that is meant for aging) involves a wide variety of sensory aspects such as color, aroma, and flavor.Physicochemical indicators, including the profiles of volatile compounds, phenolics and organic acids, and antioxidant capacity, were determined for a wide range of Cabernet Sauvignon and Merlot vintages from different areas of Bordeaux (Chira et al. 2011).Similar studies have been published on Spanish Tempranillo, Graciano, and Cabernet Sauvignon wines (Monagas et al. 2006), Chilean Cabernet Sauvignon (Lissi et al. 2014), andTreixadura wines (Vázquez-Pateiro et al. 2020).In general, these studies suggest a significant decrease in the concentration of total anthocyanins with aging and a decrease in the concentrations of catechin and proanthocyanidins, consistent with the loss of low-molecular weight phenols and the formation of higher molecular weight tannin polymers.In a more recent publication, Maioli et al. (2022) monitored the chemical and sensory properties of Sangiovese red wines after six and 12 months of aging in different types of tanks and six months in glass bottles.This study found that bottle aging enhanced chemical and sensory differences between wines aged in different types of tanks, and that they were characterized by a higher content of varietal volatiles such as norisoprenoids and terpenes.However, an exhaustive compilation of available studies on the temporal evolution of polyphenols and antioxidant activity in different wines concluded that there is no consistent pattern in the reported results (Lissi et al. 2014).
The sensory and chemical properties of young Malbec wines have been examined (King et al. 2014, Buscema and Boulton 2015, Nelson et al. 2015, Urvieta et al. 2018, 2021).Changes in polyphenol and anthocyanin profiles and elemental composition in wines from different regions of Mendoza and California were also evaluated after five years of aging (Agazzi et al. 2018).However, no previous publications have studied changes in aromatic profile during aging of Malbec wines and the effect of regionality in the aging process.This study analyzed the evolution of the volatile composition of different Malbec wines, made under identical winemaking conditions, from Mendoza and California, and compared the initial aromatic profile with that obtained after seven years of aging under controlled storage conditions.

Wine samples
Twenty-six Mendoza wines produced from grapes grown in six different districts within the departments of Luján, San Carlos, and Tupungato were studied.The study also included 15 California wines produced from grapes grown in seven districts within the counties of Napa, Sonoma, Monterey, and Yolo (Table 1; Buscema and Boulton 2015).Plots were selected to provide a representative sample of Malbec grapes from each department.The samples were harvested by hand, in uniform 500 kg batches at 24 to 25 Brix and without perceptible herbaceous notes.The trial was conducted during the 2011 harvest.Wines from Mendoza were made at the Catena Institute of Wine (Mendoza, Argentina) and the California wines were made in the pilot winery at the University of California, Davis (Davis, CA).Subsequently, the wines were stored at 16.5 ± 0.2°C in the UC Davis bottle  (Buscema and Boulton 2015).The volatile profiles of the wines were analyzed right after bottling and are identified as 't0 wines' (data reported in King et al. 2014).The same wines were then bottle-aged for seven years at 16.5 ± 0.2°C and re-analyzed (the aged wines are referred to as "t7 wines").

Volatile profile analysis
The aromatic profiles of the wines were analyzed as described (Hjelmeland et al. 2013).This method is based on the solid-phase microextraction of volatile compounds present in the headspace (HS SPME) and its subsequent analysis by gas chromatography coupled to mass spectrometry (GC-MS).Fifty-seven volatile compounds of interest were semiquantified using undecanone as the internal standard.As previously reported, these compounds are important contributors to aroma in a wide variety of red wines, including Malbec (Kotseridis et al. 2000, Campo et al. 2005, Goldner and Zamora 2007), and are associated with aroma attributes such as berries (ethyl and acetate esters), violets (ionones), herbal (C6 alcohols), sweet-caramel (phenyl acetaldehyde, linalool), and woody (oak lactone) (Escudero et al. 2007).
Chemical analyses of the identified compounds were performed in triplicate (Table 2).Chemical spectra, calculated retention times, and experimental retention times were compared to those obtained for the reference compounds in a previous work, with the exception of three compounds (vitispirane I and II and α-cedrene), due to unavailability (Hjelmeland et al. 2013).Experimental and reference retention indices and the ions selected for SIM detection are shown (Table 2).
The analysis was carried out as follows: 10 mL wine sample was combined with 50 ng L-1, 2-undecanone, and 3 g NaCl in a glass vial with magnetic crimp caps (Supelco).The samples were then exposed to a 2 cm divinylbenzene/ Chromatographic analysis was performed using an SPME inlet liner (0.7 mm i.d.; Supleco) and a DB-Wax (polyethylene glycol) capillary column (30 m, 0.25 mm i.d., and 0.25 µm film thickness; J&W Scientific).The inlet temperature was maintained at 240°C and the SPME fiber was desorbed in split mode, using a split ratio of 20:1.Helium was used as the carrier gas with a constant flow of 1 mL/min.An oven temperature gradient was used to achieve resolution of the analytes.An initial oven temperature of 40°C was held for 5 min, then increased by 3°C/min to 180°C, and then by 30°C/min to 240°C.Finally, the oven temperature was kept at 240°C for another 10 min.An electron ionization source was used, with a source temperature of 230°C and an electron energy of 70 eV.The samples were analyzed using a 6890 gas chromatograph coupled to a 5975 MSD set at 240°C (Agilent Technologies), equipped with an MPS2 autosampler (Gerstel).The instrument was controlled by Maestro (version 1.2.3.1,Gerstel) and the data were analyzed using ChemStation software (E.01.01.335,Agilent Technologies).

Statistical analyses
The factors studied were age and origin: (i) aging time, indicated by t0 (after five to six months of aging for California wines, or seven to nine months for Mendoza wines) and t7 (after seven years of aging); and (ii) geographic origin of the vineyards.This factor was examined both as the effect of region (Mendoza versus California) and of the subregions or departments (see Table 1).Principal component analysis (PCA) was used as a multivariate technique for the characterization and exploration of the samples.The confidence ellipses were calculated at a significance level of α = 0.05.Partial least squares-discriminant analysis (PLS-DA) was used to build a classification model of the samples based on their volatile profiles, using software developed by Zontov et al. (2020).

Evolution of volatile profiles
The wine volatile profiles at t0 were analyzed in a previous study (King et al. 2014).There were significant differences between the volatile compositions of Malbec wines from Mendoza and California.Mendoza wines were characterized by a greater presence of terpenes associated with floral and fruity aromas (eugenol, ionones, syringol, camphor and cymene, among others) and by volatile phenols and lactones associated with woody and spicy aromas.California wine volatile profiles were similar to each other, with a predominance of terpenes (linalool, limonene, and damascenone)   This study compares the volatile profiles of the Malbec wines at t0 with the t7 profiles, generated after seven years aging in the bottle using PCA (Figure 1).The profile of aromatic compounds in the Malbec wines changed significantly after seven years aging (confidence ellipses with a significance of α = 0.05).On the other hand, the t7 profiles clustered closer together than those of the t0 samples, demonstrating that the differences in volatile profiles decreased considerably over time.Component 1 (PC1, the abscissa) represents 98% of the explained variance.
The changes in concentrations of volatile compounds contributing to PC1 represent chemical changes in wines after seven years aging (Figure 1B, Table 3).Over time, in general there were lower concentrations of compounds associated with fruity, fresh, and floral aromas, and a simultaneous increase in the concentration of volatile compounds related to spicy, woody, and toasted aromas.This development of the so-called bouquet, bottle bouquet, or bottle-aged character, is in concordance with similar studies performed on other wine varieties (Pereira et al. 2014, Moreira et al. 2016, Cassino et al. 2019).
A recent study analyzed the effect of temperature, closure type, and aging time on the phenolic and volatile composition of Mendoza Malbec wines during 24 months of aging (Giuffrida de Esteban et al. 2019).While the aging times in that study were significantly shorter than the aging period applied in this work, similar trends were observed in the levels of different families of compounds: both studies show a decrease in the levels of terpenoids, volatile phenols, and esters over time.Similar results were found in a 20-month aging study of Pinot noir wines (Cantu et al. 2021).
Finally, it is important to note that although the wines studied here were never in contact with wood during the winemaking process, t0 wines showed the presence of numerous compounds usually associated with oak maturation, including vanillin, oak lactones, whiskey lactone, furfural, and guaiacol.Consistent with these results, descriptive sensory analysis found woody aromas in Malbec samples from Mendoza at t0, as reported by King et al. (2014) and Heymann et al. (2015).This finding demonstrates that contact with wood is not the only source of such compounds in Malbec wines, although it may be the most significant.Analysis of the same samples after seven years of aging showed that the concentrations of some of these components decreased with time.Although these findings are contrary to some reports (Morata 2019), there are no previous studies on the evolution of volatile composition in Malbec wines without oak aging and more studies are needed on this topic.A recent study evaluated aromatic profiles of Pinot noir wines with no oak exposure after eight and 20 months of bottle aging (Cantu et al. 2021).This study found no oak lactones, vanillin guaiacol, or other wood-related compounds, suggesting the phenomena observed in our study could be specific to Malbec wines.
Further data treatment was performed to assess the differences in volatile profiles between t0 and t7, and between region of origin.The full data set was divided into two subsets: a training set (70% of the samples) and a test set (the remaining 30% of the samples).The data were subjected first to mean centering.Next, the training set was used to build a PLS-DA classification model between the four classes (Mendoza_t0, California_t0, Mendoza_t7, and California_t7).Five latent variables were needed to achieve proper classification.While a high percentage of variation was explained by only two PCs in the PCA, three extra components were necessary in the PLS-DA, showing that minor components also influenced the classification.The PLS-DA model was then employed to predict the classes of the samples in the test set, achieving a successful classification of 100% of samples and demonstrating that there were significant differences in the volatile profiles of the studied samples, owing to both sample origin and bottle aging time.

Regionality after aging
Once the temporal evolution of the samples was analyzed, we compared the volatile profiles of the Malbecs  from Mendoza with those of the Malbecs from California, after seven years of aging.The analysis by PCA is shown in Figure 2A.There were significant differences in the profiles of aromatic compounds at t7 according to the region of origin.There was a greater diversity of profiles in the Mendoza samples, evidenced by the greater dispersion of the sample scores in the PCA graph.In this analysis, PC1 represented 99% of the explained variance.The individual chemicals contributing to PC1 were also determined (Figure 2B).
The more significant contributions to the volatile profiles of the analyzed wines at t7 are presented (Table 4).Wines from Mendoza were characterized by many compounds of interest, associated with a broad variety of aromas such as fruity and fresh, spicy, woody, and vanilla (Peinado et al. 2004, Escudero et al. 2007, Morata 2019).For their part, California wines were characterized by less complex aromatic profiles, with compounds associated with fruity, spicy, and woody descriptors.Some characteristic compounds were present in wines from both origins, but most of these were more abundant in Argentinean Malbecs.While the difference in aromatic diversity observed in this study is mostly associated with regionality, the aromatic profiles could also have been influenced by the California vines being grafted, while those from Argentina were own-rooted.Rootstock has been reported to have an impact on the volatile composition of wines (Romero et al. 2019, Wang et al. 2019, Vilanova et al. 2021).
Further analysis of the volatile compositions of the seven-year-old wines showed that there was no differentiation by subregion or department.As previously mentioned, the same results were obtained when analyzing the aromatic data at t0, i.e., no particular trends or clustering by department or subregion was observed.However, it is important to point out that our previous studies have been able to discriminate these and other similar Malbec samples from different departments, using descriptive sensory analysis and polyphenolic profiles, in both young and aged Malbec samples (Agazzi et al. 2018, Urvieta et al. 2018, 2021).This shows there is a significant effect of season and/or site on sensory perception and chemical composition of the same set of wines.
Finally, ionones, a group of compounds of particular importance in Malbec that are associated with violet aroma, was recorded at t0 in the wines from both regions, but only the Mendoza samples showed detectable concentrations at t7.These compounds not only are associated with desirable floral aromas, but also interact with other volatile components, enhancing and/or masking the perception of other aromas (Escudero et al. 2007, Ferreira et al. 2016).Peinado et al. 2004, Escudero et al. 2007, and Morata 2018.b Significantly different concentration between one region and the other.
Since the aroma compounds were only semiquantified, their absolute concentrations are unknown and cannot be compared with the thresholds of perception.However, the presence of ionones and other compounds of interest (furfural, camphor, vanillin, eugenol, etc.) after seven years of aging shows that the Malbec wines maintained a rich volatile profile, characteristic of their region of origin, suggesting high AP.
The authors propose to continue this study by evaluating the contribution of volatile compounds to aged Malbec wine aroma using gas chromatography coupled with olfactometry (Welke et al. 2021) and descriptive sensory analysis.However, our current findings could have a direct impact on decision making for wineries in Mendoza and California.For example, the shorter lifespan of desirable volatile compounds in California Malbecs suggests that California wineries could benefit from using less oak and selling their wines more rapidly to preserve a distinctive volatile fingerprint.California Malbecs could be used as a blender if a longer AP is desired.In contrast, this study indicates that Malbec wines from Mendoza could be used both as a blender and as a single varietal.Finally, these results should encourage California growers to try different plant materials to search for Malbec wines that better preserve their unique volatile profiles.

Conclusions
The evolution of aromatic compounds during a relatively long period of bottle-aging (i.e., seven years) of Malbec wines was examined for the first time.When comparing the results obtained at the beginning and after seven years of aging, it was observed that there is a significant time effect on the volatile profiles of Malbec wines.Several compounds associated primarily with fruity and floral aromas are predominant in the younger wines, while the t7 wines are characterized by an abundance of compounds related to woody, smoky, fresh, vanilla, and earthy aromas.Even after seven years of aging, the wines are easily differentiated by region of origin.Although a loss of diversity is observed in the aromatic profiles over time, it is not very significant in Mendoza wines.Furthermore, a large variety of aromatic compounds considered 'desirable' or 'of interest' were present in the Mendoza Malbecs after seven years of bottle aging.Thus, the Malbec wines from Mendoza were more suitable for aging than the wines from California.The results obtained in this study could prove helpful to wineries when making decisions about winemaking, aging, and commercialization of Malbec wines.

Figure 1 A)
Figure 1 A) Principal component analysis (PCA) separation of samples by aging time and region using volatile composition.Mendoza_t0 are shown as pink dots; California_t0, as cyan dots; Mendoza_t7, as pink diamonds; and California_t7, as cyan diamonds.Confidence ellipses were calculated at a significance level of α = 0.05.B) Percentage contribution of variables (volatile compounds) to PC1 in PCA of young versus old wine samples.

Figure 2
Figure 2 (A) Principal component analysis (PCA) separation of samples by region of origin at t7, using volatile composition.Argentinian wines are shown as pink dots and California wines, as cyan dots.Confidence ellipses were calculated at a significance level of α = 0.05.B) Percentage contribution of variables (volatile compounds) to PC1 in PCA comparing wine regions (Mendoza versus California) after seven years of aging.

Table 1
Wine regions from Mendoza, Argentina and California, USA included in the study.

Table 2
A list of the compounds measured using headspace solid-phase microextraction gas chromatography-mass spectrometry, their CAS number, retention time, retention index (RI), calculated retention index (CRI), and selected ion monitoring (SIM) qualifying ions.

Table 2 continued
A list of the compounds measured using headspace solid-phase microextraction gas chromatography-mass spec- trometry, their CAS number, retention time, retention index (RI), calculated retention index (CRI), and selected ion monitoring (SIM) qualifying ions.a Acree and Arn 2004.b El-Sayed 2023.c Humpf and Schreier 1991.

Table 3
Predominant volatile compounds at t0 and t7 in Malbec wines from Mendoza and California, with origin and associated aromatic descriptors.several esters.Multivariate analysis showed that it is possible to differentiate Mendoza wines from California wines based on volatile composition.However, the wines could not be separated more finely by department/county of origin.

Table 4
Predominant volatile compounds at t7 in Malbec wines from Mendoza and California, and associated aromas.