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Toasting. Ellagitannins. Volatile composition. Sensory analysis. ABSTRACT. Malolactic fermentation (MLF) and oak barrels aging are two oenological processes ...

Food Chemistry xxx (2018) xxx-xxx

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Food Chemistry


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Use of oak wood during malolactic fermentation and ageing: Impact on chardonnay wine character M.R. González-Centenoa⁠ ,⁠ b⁠ ,⁠ c⁠ , K. Chiraa⁠ ,⁠ b⁠ ,⁠ c⁠ , P.L. Teissedrea⁠ ,⁠ b⁠ a b c

Univ. Bordeaux, ISVV, EA 4577, Enologie, 210 Chemin de Leysotte, 33140 Villenave d’Ornon, France INRA, ISVV, USC 1366 Enologie, 210 Chemin de Leysotte, 33140 Villenave d’Ornon, France Tonnellerie Nadalié, 99 Rue Lafont, 33290 Ludon-Médoc, France



ARTICLE INFO Keywords: Malolactic fermentation Oak wood barrel Toasting Ellagitannins Volatile composition Sensory analysis

Malolactic fermentation (MLF) and oak barrels aging are two oenological processes which modify wine composition and sensory characteristics. The effect of MLF-container (stainless steel tanks, barrels) and barrel toasting (T1, T2, T3) on ellagitannin concentration, volatile composition and sensory attributes of Chardonnay wines was evaluated. Barrel toasting had higher impact on ellagitannin content than MLF-container. When comparing both MLF-modalities, barrel-fermented wines exhibited greater amounts of vanillin and whiskey-lactones but lower concentrations of fruity aroma compounds. Regarding sensory analysis, greater citrus and floral aromas were perceived for MLF-tank wines, and higher nuts aroma for MLF-barrel wines. Using barrels as MLF-containers i) did not change the aroma perception defining Chardonnay character (peach, apricot and gun flint); ii) did not impact the aromatic intensity and persistence, sweetness, acidity, mouthfeel volume and/or bitterness; and iii) did not confer significantly higher overall woody aromas which might mask fruity, floral and mineral (gun flint) character.

1. Introduction

Chardonnay is one of the oldest and most widely distributed wine grape varieties in the world. It readily adapts to a myriad of divergent terroirs in nearly every winegrowing region. This vast flexibility leads to production of countless Chardonnay wine styles, with particular aroma and flavor nuances, and different levels of sweetness, bitterness and acidity (Gambetta, Bastian, Cozzolino, & Jeffery, 2014). Due to its ageing ability and its ease in reflecting the area where it is grown, Chardonnay variety has gained mass popularity. In fact, no other white grape variety allows winemakers to produce wines with such complexity. Depending on the targeted style, a wide range of processes made during winemaking may shape the overall aroma and flavor of the final Chardonnay wine (Gambetta et al., 2014). Seeking for a touch of uniqueness in an increasingly globalised wine market, malolactic fermentation (MLF) of Chardonnay wines, as well as their ageing in oak barrels, has become a common practice in some winegrowing regions, despite both techniques being typical of red vinification.

During MLF, lactic acid bacteria (LAB) not only contribute to deacidify and stabilize the wine, by transforming the malic acid into lactic acid, but may also enhance the aroma and flavor complexity of the resulting wine, by producing some odor-active compounds and/or transforming both grape and yeast derived volatiles and flavor precursors related to its typicity (Antalick, Perello, & de Revel, 2012). Ageing in oak barrels provides wine with a slow but constant micro-oxygenation process, which may help to stabilize wine color, and with different volatile and non-volatile wood compounds, which may impact its final organoleptic perception (Liberatore, Pati, Del Nobile, & La Notte, 2010). The principal factors affecting the release of these compounds (mainly ellagitannins and woody volatiles) into wine are the composition of the wine matured in barrels, the ageing length and the available amount of wood compounds potentially extractable, which, in turn, is especially conditioned by the barrel toasting (Chira & Teissedre, 2015; González-Centeno, Chira, & Teissedre, 2016). On the whole, other than the direct grape contributors to wine aromatic bouquet, an array of volatile and non-volatile components, which also contribute to the character, style and overall quality of Chardon

Email address: [email protected] (K. Chira) https://doi.org/10.1016/j.foodchem.2018.11.049 Received 25 May 2018; Received in revised form 9 November 2018; Accepted 9 November 2018 Available online xxx 0308-8146/ © 2018.

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the Tonnellerie Nadalié (Ludon-Médoc, France) wood yard. Once assembled, barrels (225 L) were submitted to different toasting procedures using the traditional way over an oak wood fire. Three toasting levels were provided (T1, T2, T3), all of them submitted to a pre-toasting step of 20 min at 45 °C. For the T1 and T2 toastings, a gradual rise in temperature up to 55 ± 2 °C and 52 ± 2 °C, respectively, was applied for 36 min. In the case of T3 toasting, temperature was gently increased up to 60 ± 2 °C for 10 min, and then, gradually decreased down to 45 ± 2 °C for 10 min more. The barrel heads were not toasted. For the purpose of the study, five barrels of each toasting procedure were provided to the wine cellar.


nay wines, may arise during malolactic fermentation and barrel ageing steps (Gambetta, Cozzolino, Bastian, & Jeffery, 2016). Despite that, there is still scarce scientific research focused on the use of these red vinification techniques for white wine. Specifically, Liberatore et al. (2010) and Herrero et al. (2016) have recently studied the influence of ageing in oak wood barrels (10 and 12 months, respectively) on the volatile composition of Chardonnay wines (fruity and/or woody aromas). Liberatore et al. (2010) demonstrated that performing fermentation and ageing in barrels with lees, the content of flavor active compounds such as lactones, terpenoids and esters, among others, was enhanced, and taste was improved, in comparison to fermentation and ageing in stainless steel tanks or in barrels with partial removal of lees. Herrero et al. (2016) observed that nearly all the studied woody volatiles increased with the ageing time, and specifically for furfural, guaiacol and vanillin derivatives, their contents raised with the intensity of the toasting level. To our knowledge, there are no studies evaluating the impact of MLF-container on the phenolic, aromatic and sensory profiles of Chardonnay wines, and/or regarding the effect of other barrel toastings than the common ones. Thus, a more in-depth understanding of the impact of both malolactic fermentation and barrel ageing steps on Chardonnay character is paramount for wine producers. Within this context, the aim of the present research was to further investigate the effect of both MLF-container and barrel toasting on the phenolic composition (including ellagitannins), the aromatic profile and the sensory attributes of two sets of Chardonnay white wines: one in which MLF and ageing were carried out in oak barrels, and another where MLF was performed in stainless steel tanks previous to barrel ageing.

2.2. White wine vinification and sample collection


Chardonnay grapes (Vitis vinifera L.) were manually harvested at maturity in Domaine Costa Lazaridi winery (Adriani, Drama, Greece) during the 2013 vintage. Grapes were crushed and destemmed the day of harvest (Enoitalia WE223S, Italy). A pneumatic press (Vaslin–Bucher RPS 50, France) was used filled at 75–80% of its capacity. Potassium metabisulphite (4.5 ± 0.5 g/hL) was added during the transfer of must to stainless steel tank (maximum capacity 40 hL). Commercial active dry yeast of type Saccharomyces cerevisiae (Zymaflore VL1, Laffort) was included at a rate of 25 g/hL to perform alcoholic fermentation at 18 – 20 °C. Two sets of experiments were performed depending on the container where malolactic fermentation (MLF) took place (Fig. 1). Thus, when alcoholic fermentation concluded, wine was partly transferred to new oak barrels presenting T1, T2 and T3 toastings (a total of six barrels of 225 L were filled, two barrels per toasting) to perform malolactic fermentation (MLF-barrel modality). Meanwhile, the rest of the wine remained in the same initial stainless steel tank of variable capacity to conduct MLF (MLF-tank modality). In both types of container, wine was inoculated with lactic acid bacteria (Lactoenos B7 Direct, Laffort) at a rate of 30 g/ hL, and MLF extended for two months at a maintained temperature of 18 – 20 °C. Once the MLF was finished (malic acid content ≤0.2 g/L, controlled every two days by an enzymatic test kit), wines that went through MLF

2. Materials and methods

2.1. Oak wood origin and drying conditions

All barrels used were made up of French oak from two species (95% Quercus petraea and 5% Quercus robur) from the same forest located in the Center region of France. The raw staves (100 cm × 11 cm × 2.2 cm) were naturally seasoned for 24 months in

Fig. 1. Experimental design of the winemaking process followed in the present study. 2

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specified by González-Centeno et al. (2016). Retention times and mass spectra of the pure reference standards were used to identify all target compounds. Results were calculated from calibration curves previously established using those pure reference standards analyzed under the same conditions than wine samples. A duplicate per barrel was performed (n = 4 per each MLF-container × toasting).


in barrels were immediately racked and, after barrel cleaning and sanitization, were returned to the same ones. Potassium metabisulphite was then added at 6 g/hL. Wines that went through MLF in stainless steel tanks, were racked and transferred to new oak barrels (a total of six barrels of 225 L were filled, two barrels per toasting) presenting the same three toastings than the MLF–barrel modality. Potassium metabisulphite (6 g/hL) was also added to the wine of the MLF-tank modality. From the five barrels of each toasting procedure provided to the wine cellar, two were used for the wines of the MLF-tank modality and another two for the wines of the MLF-barrel modality. The fifth one was used for the ullage or fill level of the barrels during ageing. Wines were kept in oak barrels for ageing during 12 months at a controlled temperature of 14 – 16 °C. All three toasting methods (T1, T2, T3) described above were tested in each set of experiments. After 12 months of wood contact, wines from oak barrels of each MLF-modality and toasting method were bottled, and then, directly sent to the laboratory at 16 °C for further analysis.

2.6.1. Woody aroma For the identification of the target compounds, selected ions were m/z 151 for vanillin; m/z 164 for eugenol and isoeugenol; m/z 124 for guaiacol; m/z 99 for β-methyl-γ-octalactone; m/z 96 for furfural; m/z 110 for 5-methylfurfural; and m/z 83 for the internal standard (dodecan-1-ol). 2.6.2. Fruity aroma The following ions were used to identify the target compounds: 2-phenylethyl acetate, m/z 104; ethyl 2-methylbutanoate, m/z 102; ethyl butyrate, ethyl hexanoate, ethyl octanoate, ethyl decanoate, ethyl isobutyrate and ethyl 3-methylbutanoate, m/z 88; isoamyl acetate, m/z 70; and m/z 56 for isobutyl acetate, butyl acetate and hexyl acetate.

2.3. Oenological parameters in wines Conventional oenological parameters of wines such as pH, alcoholic strength (% vol.), titratable and volatile acidity (g/L tartaric acid), were determined by Infrared Spectrometry with Fourier Transformation (IRTF) with a WineScan™ Flex (FOSS Analytical, Denmark), which was previously calibrated with wine samples analyzed in accordance with official OIV methods (2016). All parameters were determined in duplicate per barrel (n = 4 per each MLF-container × toasting).

2.7. Sensory analysis


All evaluations were conducted in a standard sensory-analysis chamber (ISO-8589, 1988), equipped with separate booths, where an uniform source of lighting, absence of noise and distracting stimuli were guaranteed, and the ambient temperature was maintained at 19 – 22 °C. Standard black wine glasses (ISO-3591, 1997) were used for both training and tasting sessions, covered with a glass Petri dish to minimize the escape of volatile components and randomly coded with three-digit numbers. The position of the wine samples was balanced in all sensory tests. To assess the impact of the MLF-container and the barrel toasting method on the character of Chardonnay wine, sensory analyses were performed by a panel of 19 expert judges (sixteen women and three men). All of them had a wine tasting degree (DNO, Graduate Diploma in Oenology or DUAD, a professional tasting diploma) and were staff at the Institute of Vine and Wine Sciences of the University of Bordeaux (France). Chardonnay character for this research was defined according to the odorant attributes of higher occurrence (yellow fruits, citrus, white flowers, nuts, gun flint) in the sensory profiling of Chardonnay wines described by Gros et al. (2017). Olfactory and gustative panel training was first performed to familiarize the subjects with Chardonnay character, aroma recognition and wine taste. Secondly, descriptive sensory analysis was conducted to assess the sensory profile of wines matured in barrels with different toasting methods, for each MLF-modality separately (two formal tasting sessions per MLF-modality). Thirdly, for each barrel toasting method, discriminatory tests including triangle test (ISO-4120, 2004) and bilateral paired comparison test (ISO-5495, 2005) were carried out to evaluate whether the panel was able to distinguish between wines that went through MLF in barrels or in stainless steel tanks, and if yes, to identify to what sensory descriptors they relate that difference (two formal tasting sessions per each barrel toasting method). The olfactory and gustative preference was requested to the trained judges at the end of all tasting sessions. The olfactory training consisted of three formal sessions where the panel was asked to identify the main aromas of the defined Chardonnay character, as well as the ‘overall woody’ descriptor, used to describe all olfactory sensations brought about by the oak wood (vanilla, smoky, toasty). For this purpose, fresh daily reference standards of peach and apricot (yellow fruits), of lemon, bergamot orange and grapefruit (citrus fruits), of honeysuckle and jasmine (white flowers), and of almond and hazelnut (nuts), were prepared. Dried leaves of ver

2.4. Total phenolic content

Total phenolic content of Chardonnay wines was spectrophotometrically determined, by using an automated microplate reader (FLUOstar Optima, BMG LabTech, France). Experimental procedure was as previously reported by González-Centeno et al. (2016), but wines were diluted at a ratio 1:4 with distilled water. Measurement was performed in triplicate per barrel (n = 6 per each MLF-container × toasting). 2.5. Oak ellagitannins of wines: extraction and identification by HPLC

Ellagitannin fraction was obtained after column fractionation as described by González-Centeno et al. (2016). Prior to HPLC–UV/MS analysis, the solid residue obtained was dissolved in H2⁠ O/HCOOH (996/ 4, 1 mL) and filtered through a syringe (PTFE filters, 0.45 μm of pore size). Ellagitannin identification was performed on a reversed-phase LiChrospher 100 RP18 (250 mm × 4.6 mm, 5 μm) column by using a Thermo-Finnigan Surveyor HPLC system. Elution conditions, flow rate, and composition of the mobile phases were adapted from Michel et al. (2011). Each target compound (castalagin, vescalagin, grandinin and roburins A-E) was quantified by using their molecular ion. Results, expressed as equivalents of castalagin (mg castalagin equivalents/L wine), were calculated relative to the response of the chlorogenic acid (20 mg/ L) used as an internal standard. All ellagitannins analyses were performed in duplicate per barrel (n = 4 per each MLF-container × toasting). 2.6. Volatile composition of wines: extraction and gas chromatography analysis

Woody and fruity aroma profiles were determined according to the gas chromatography methods described by Barbe and Bertrand (1996), and Antalick, Perello, and de Revel (2010), respectively. Experimental procedures of the volatile extraction prior to gas-chromatographic analyses, equipment and calibration conditions were considered as


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analysis, the Scheirer-Ray-Hare test, when residuals were not distributed normally and/or data presented heterogeneity in variance. Sensory results were based on the probability theory that the number of right answers follows a binomial distribution (n, p = 1/3 for triangle test, and p = 1/2 for bilateral paired comparison test), where n is the panel size (n = 20). Wines from both MLF-modalities were considered as differently perceived for a probability lower than 5%.


bena were used for aromatic plants, and firecracker for the mineral nuance of gun flint. Only during the first training session, a reference per aroma was smelt previous to assessors’ blind evaluation of all reference standards. At the end of the olfactory training sessions, all participants were homogenized, correctly identified all aromas defining Chardonnay character, or at least all the corresponding aroma families, and became familiar with the olfactory descriptors evaluated in this research. From a gustatory point of view, judges were also trained to identify and scale of sweetness and bitterness, as previously described by Chira and Teissedre (2013). For the descriptive analysis, a ten-point scale (0 = ‘non-existent’, 10 = ‘at maximum intensity’) was implemented for scoring of the sensory attributes. Judges were asked to first evaluate the orthonasal odour (aromatic intensity, overall woody, peach and apricot flavours, citrus fruits, nuts, floral and aromatic plants, flint), and then, after a short break, the wine taste (aromatic persistence, sweetness, acidity, mouthfeel volume, bitterness). The intensity level of each descriptor was then expressed as the mean value of all the judges. At the end of the tasting session, the olfactory and gustative preference among the three toasting methods was also requested for each MLF-modality. For the triangle test, three glasses were presented and judges were asked to indicate the one olfactory perceived as different from the others. For the bilateral paired comparison test, judges started with evaluation of the orthonasal odor (aromatic intensity, overall woody, peach and apricot flavors, citrus fruits, nuts, floral and aromatic plants, flint), and then, after a short break, the taste (aromatic persistence, sweetness, acidity, mouthfeel volume, bitterness). At the end of the tasting session, the olfactory and gustative preference between both MLF-modalities was also requested for each barrel toasting method.

3. Results and discussion

3.1. Oenological parameters and total phenolics of wines


Regardless of the MLF-container and the barrel toasting, all Chardonnay wines presented a pH of 3.6 ± 0.1, an alcohol strength of 14.3 ± 0.0% vol., a titratable acidity of 3.5 ± 0.2 g tartaric acid/L wine and a volatile acidity of 0.5 ± 0.1 g tartaric acid/L wine. It is well known that pH, alcohol concentration and wine matrix may condition the rate of MLF (Gockowiak & Henschke, 2003). Nevertheless, in the present research, the same initial wine was considered for both sets of experiments and both pH and alcohol concentration values were the same for all final wines. Thus, only the different wine evolution according to the MLF-container and the barrel toasting method may justify the significant differences observed among their phenolic, aromatic, and sensory attributes. With regard to the total phenolic content, only Chardonnay wine aged in T2 barrels presented significant differences (p < 0.05) between both MLF modalities (346 ± 3 and 379 ± 9 mg gallic acid equivalents/ L wine, respectively, for MLF-tank and MLF-barrel). In this case, as previously observed for red wines (González-Centeno, Chira, & Teissedre, 2017; Vivas, Lonvaud-Funel, Glories, & Augustin, 1995), the barrel-fermented wine showed a slightly higher total phenolic content than the corresponding tank-fermented one. In contrast, regarding the toasting effect and more particularly in the case of T1 and T3 toastings, total phenolics of both MLF modalities did not differ significantly (p > 0.05).

2.8. Statistical analysis

All experimental results were presented as mean values with their corresponding standard deviations. Statistical analysis was carried out using the statistical package R version 3.1.1 (R Foundation for Statistical Computing, Wien, Austria). Normality and homocedasticity of the residuals were evaluated for all parameters, by using the Shapiro–Wilk test and Levene’s test, respectively. When residuals were normally distributed and populations presented homogeneity in variance, parametric tests were used. Thus, data was submitted to two-way ANOVA (MLF-container × toasting method). If the interaction p-value was statistically significant, Tukey test was directly applied to evaluate the degree of the significant differences. If not, previously to the Tukey evaluation, a one-way ANOVA was run individually for data corresponding to each significant factor. Differences at p ≤ 0.05 were considered to be statistically significant. Two-way ANOVA was substituted by the corresponding nonparametric

3.2. Ellagitannin composition of wines The individual ellagitannin composition of Chardonnay wines investigated is described in a detailed form in Table 1. To the best of the authors’ knowledge, no studies addressing the comparison of the oak wood ellagitannin content in white wines depending on the MLF container have been previously published in the literature. Total ellagitannin content of Chardonnay wines after 12 months of oak barrel ageing, calculated by adding up the individual concentration of each ellagitannin compound, varied from 7.8 ± 0.2 to 17.4 ± 1.3 mg castalagin equivalents/L wine for T2-tank and T2-barrel, respectively. Since no data for the ellagitannin individual composi

Table 1 Ellagitannin composition (n = 4) of Chardonnay wines of the two MLF modalities (in stainless steel tanks and in barrels) after 12 months of ageing in oak barrels representing different toasting processes (T1, T2, T3). T1 toasting

Roburin A Roburin D Vescalagin Castalagin Roburins B + C Gradinin Roburin E Total ellagitannins

T2 toasting

T3 toasting

MLF in tank

MLF in barrel

MLF in tank

MLF in barrel

MLF in tank

MLF in barrel

0,04 ± 0,01b 0,08 ± 0,00b 1,95 ± 0,01 a 6,31 ± 0,42b 0,17 ± 0,00b 2,60 ± 0,03 a 2,76 ± 0,09b 13,92 ± 0,43b

0,04 ± 0,00b 0,06 ± 0,00c 1,49 ± 0,00b 5,18 ± 0,55b 0,20 ± 0,03b 2,52 ± 0,29 a 2,48 ± 0,01b 11,97 ± 0,62c

0,04 ± 0,00b 0,06 ± 0,00c 0,88 ± 0,08 d 3,18 ± 0,13c 0,19 ± 0,05b 1,71 ± 0,15b 1,80 ± 0,03c 7,85 ± 0,22 d

0,07 ± 0,00 a 0,12 ± 0,02 a 2,05 ± 0,08 a 8,56 ± 1,29 a 0,37 ± 0,04 a 2,40 ± 0,26 a 3,78 ± 0,24 a 17,35 ± 1,34 a

0,05 ± 0,00b 0,08 ± 0,01b 1,40 ± 0,11 bc 4,98 ± 0,50b 0,25 ± 0,03b 2,48 ± 0,39 a 2,40 ± 0,27b 11,65 ± 0,70c

0,05 ± 0,00b 0,10 ± 0,00 a 1,25 ± 0,02c 4,56 ± 0,25 bc 0,23 ± 0,01b 2,29 ± 0,06 ab 2,48 ± 0,08b 10,96 ± 0,27c

All results were expressed in mg castalagin equivalents/L wine. T, toasting. MLF, malolactic fermentation. Lower case letters a–d show significant differences between categories MLF-container × toasting method for each ellagitannin (p < 0,05). 4

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barrel toasting process and/or formed as a consequence of the wood-wine interaction. Among them, furfural and 5-methylfurfural, cisand trans-whiskey lactones, guaiacol, eugenol and vanillin, have been described by Gambetta et al. (2014) as odorants derived from oak wood contact, or formed during ageing, that are important to the typicity of Chardonnay wines. In the present research, the impact of MLF-container on these woody volatiles of 12-months barrel-aged Chardonnay wines was investigated for three different toasting methods (Fig. 2). Except for 5-methylfurfural, the two-way ANOVA of the raw experimental data disclosed that the MLF-container and the toasting method, as well as the interaction between both factors, significantly impact (p < 0.05) the oak wood volatiles concentration of the studied wines. The total content of woody volatiles in Chardonnay wines, calculated by adding up the individual concentration of each above-mentioned compound, ranged from 3303 to 4207 μg/L for MLF-barrel wines, and from 6603 to 9326 μg/L for MLF-tank wines. For both MLF-modalities, T1 toasting led to the highest values, and T3 toasting to the lowest ones. Unlike the expected behavior, for all three toasting methods, total woody volatiles content was from 2.0 to 2.2-fold greater in MLF-tank wines than in wines with oak wood contact during MLF (p < 0.05). It is well known that furanic compounds have low direct impact on the organoleptic attributes of wines due to their high perception thresholds (furfural at 20 mg/L and 5-methylfurfural at 45 mg/L) (Boidron, Chatonnet, & Pons, 1988). Thus, if only the main direct contributors to the overall oak wood aroma (whiskeylactones, eugenol, vanillin) are considered for the total woody volatile content, higher values were observed in MLF-barrel modality (by 14.0 – 50.7% depending on the toasting), confirming the behavior previously stated in the literature for Cabernet Sauvignon red wine (González-Centeno et al., 2017).



tion in white wines exists in the literature, only a comparison with red wines can be made. Indeed the values found are in agreement with the order of magnitude previously reported in the literature for the major ellagitannin compounds in red wines (Glabasnia & Hofmann, 2006; González-Centeno et al., 2016, 2017; Jourdes, Michel, Saucier, Quideau, & Teissedre, 2011; Michel et al., 2011, 2013). Regardless of the MLF-container and/or the barrel toasting, castalagin was the major ellagitannin with a contribution between 40 and 49% to the total content. Meanwhile, the dimers roburin A and D, as well as the glucosidic dimers B and C, were present as minor constituents accounting for less than 2.5%, in all cases. Two-way ANOVA results revealed that barrel toasting defines the presence or absence of significant differences between both MLF-modalities with regard to the wine ellagitannin content. Two different behaviors were observed. In the case of T1 and T3 toastings, MLF-tank wines presented similar values of individual and total ellagitannins compared with the corresponding MLF-barrel wines (p > 0.05). Nevertheless, for the T2 toasting, barrel-fermented wine presented between 1.4and 2.7-fold greater contents of individual ellagitannins than wines having undergone MLF in tanks. These observations suggest that the lower temperature of T2 toasting compared to those of T1 and T3 toastings might influence not only the available amount of ellagitannins present in barrels, but also their extractability when MLF is performed in these oak wood vessels. 3.3. Volatile composition of wines

3.3.1. Woody aroma Overall woody aroma of barrel-aged wines results from different chemical compounds naturally present in oak wood, originated in the

Fig. 2. Woody volatile composition (n = 4) of Chardonnay wines of the two MLF modalities (in stainless steel tanks and in barrels) after 12 months of ageing in oak barrels representing different toasting processes (T1, T2, T3): (A) furfural; (B) 5-methylfurfural; (C) cis-whiskey lactone; (D) trans-whiskey lactone; (E) eugenol; (F) guaiacol; (G) vanillin. MLF, malolactic fermentation. If interaction between both factors (MLF-container and toasting) was not significant at the two-way ANOVA, one-way ANOVA was run for each factor separately: then, lower case letters x and y show significant differences between toasting methods for each MLF container modality, and * show significant differences between MLF container modalities for each toasting method (p < 0.05). If interaction between both factors (MLF-container and toasting) was significant at the two-way ANOVA: lower case letters a–d show significant differences between categories MLF-container × toasting method for each volatile compound (p < 0.05). 5

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12-months in MTAA (medium toast with watering) barrels after undergoing tank/barrel MLF (González-Centeno et al., 2017). In contrast, wines from T2 barrels presented a similar proportion of both isomers (ratio cis/trans of 1.2) for both MLF-modalities. Regardless of the toasting method, wines in which MLF occurred in barrels presented greater vanillin concentration (by 16.8 – 41.6% depending on the toasting) than the corresponding tank-fermented wines (p < 0.05). These results indicate the capacity of LAB to convert vanillin glycoside precursors present in oak wood, enhancing the vanilla flavor of the resulting wine (Bloem et al., 2008; Bloem, Lonvaud, Bertrand, & de Revel, 2006; de Revel et al., 2005). It is worth noting that, among the three barrel toastings, T3-barrel wines exhibited the lowest cis-whiskeylactone values, and T2–barrel wines showed the highest trans-whiskeylactone content for both MLF-modalities, as well as the greatest guaiacol and the lowest vanillin levels for tank-fermented modality. Meanwhile, T1 toasting led to the greatest guaiacol and vanillin contents in MLF-barrel wines, and the lowest levels of oak lactones, guaiacol and eugenol in tank-fermented wines (p < 0.05). As observed, each toasting method displayed a particular woody aroma profile, also dependent of the MLF-container. On the whole, the woody aroma is more influenced by the MLF modality than the toasting method. This may be attributed to the close toasting temperatures used for the experimental design.



In terms of quantification of the individual compounds (whiskeylactones, eugenol, vanillin), a particular woody aroma profile was established for each Chardonnay wine considered (p < 0.05). Nevertheless, a general trend of distribution persisted throughout all them, regardless of the MLF-container and/or the barrel toasting. Specifically, among the main contributors to the oak wood aroma, vanillin was the major woody volatile (302 – 488 μg/L wine), found at concentrations above its perception threshold (400 μg/L) (Boidron et al., 1988) for wines fermented in barrels with T1 and T3 toastings. The cis-whiskeylactone was the second main component (262 – 380 μg/L), closely followed by trans-whiskeylactone (168 – 320 μg/L). It is noteworthy to mention that both oak lactones were present at above-thresholds levels (20 and 140 μg/L for cis- and trans- isomers, respectively) (Brown, Sefton, Taylor, & Elsey, 2006) in all wine samples. Herrero et al. (2016), who studied Chardonnay wines after 12 months of barrel ageing, also observed whiskeylactone amounts above their perception threshold for all wines and vanillin levels greater than its threshold for half of the wines. Liberatore et al. (2010) described the same behaviour for whiskeylactones after 10 months of barrel ageing. Navarro et al. (2018) studied the woody volatile content of a discoloured Macabeo white wine aged for 12 months in French oak barrels with different toasting levels and reported values of the same order of magnitude as those observed for Chardonnay in the present research. Regardless of the toasting method, tank-fermented wines presented significantly higher contents of furfural and 5-methylfurfural than the corresponding MLF-barrel wines (p < 0.05), ranging from a 2.3 to a 2.8-fold increase. This phenomenon, attributed to a potential LAB capacity to biodegrade these furanic compounds to their corresponding alcohols during MLF in barrels (Chatonnet, Dubourdieu, & Boidron, 1991; Hernández-Orte et al., 2009), has been previously reported for Cabernet Franc (Izquierdo-Cañas, Mena-Morales, & García-Romero, 2016) and Cabernet Sauvignon (González-Centeno et al., 2017) red wines, but never for the Chardonnay wine. With regard to the barrel toasting, only wines from T1 barrels significantly differed (p < 0.05) from those of T2 and T3 barrels, displaying greater values of both furfural (MLF-tank modality) and 5-methylfurfural (both MLF-modalities). These last results suggest that maintaining the toasting temperature at 55 °C (in the case of T1 toasting) enhances the concentration of furanic compounds. By comparing both MLF-modalities of T1 toasting, higher contents of guaiacol, oak whiskey lactones and eugenol were found when MLF occurred in barrels rather than in tanks (p < 0.05): specifically, 44% more for eugenol, 75% more for guaiacol, and 25% and 53% more for isomers cis and trans of oak whiskey lactones, respectively. Even though differences were not statistically significant (p > 0.05), the same general trend was also detected for T2 and T3 toastings. This enhancement, previously noted in the literature for red wines after tank/barrel MLF (de Revel, Bloem, Augustin, Lonvaud-Funel, & Bertrand, 2005; Hernández-Orte et al., 2009; Izquierdo-Cañas et al., 2016), may be attributed to the LAB capacity to interact with wood components by the action of glycosidases, releasing additional quantities of certain woody volatiles into wine (Bloem, Lonvaud-Funel, & de Revel, 2008). Taking into account that the same LAB strain was used for all three toasting methods, and that the greater extraction of those compounds was only relevant for T1 toasting, it is suggested that the barrel toasting method may play an important role on the availability of their corresponding precursors in oak wood, by directly impacting on their formation rate during the toasting process or on the access of LAB to interact with them. Both MLF-container and barrel toasting method affected the ratio between the two isomers of oak lactones. The cis/trans ratio of the barrel-fermented wines were 1.4 and 1.1 for T1 and T3 toastings, while MLF-tank wines increased this value to 1.7 and 1.5, respectively. The same trend was described for Cabernet Sauvignon wines aged during

3.3.2. Fruity aroma A large number of volatiles, belonging to different chemical families such as alcohols, C1⁠ 3-norisoprenoids, esters, thiols, lactones, terpenes and acids, are detected in the headspace of Chardonnay wines. Among them, esters, which contribute to the fruity character of wines, are one of the predominant chemical groups (Welke, Zanus, Lazzarotto, & Alcaraz Zini, 2014). In the present research, the impact of MLF-container on the Chardonnay wine concentrations of ethyl esters of straight-chain fatty acids (ethyl butyrate, ethyl hexanoate, ethyl octanoate, ethyl decanoate), higher alcohol acetates (isoamyl acetate, isobutyl acetate, butyl acetate, hexyl acetate, 2-phenylethyl acetate) and ethyl esters branched acids (ethyl isobutyrate, ethyl 2-methylbutanoate, ethyl 3-methylbutanoate) was investigated (Table 2). Individual fruity volatile concentrations obtained in the present research were in agreement with the bibliographic ranges reported by Gambetta et al. (2014) for Chardonnay wines, except for the lower hexyl acetate and the higher ethyl decanoate and ethyl 2-methylbutanoate contents. Among the ethyl esters of straight-chain fatty acids, ethyl octanoate (1897–2061 μg/L wine), ethyl decanoate (1015–1286 μg/L wine) and ethyl hexanoate (1109–1163 μg/L wine) were the predominant ones, in that order. All of them exhibited concentrations above their olfactory perception thresholds, providing the wine with pear, apple, pineapple and/or floral notes. Isoamyl acetate, characterized by banana flavor, was the main component among the higher alcohol acetates, with values ranging from 707 to 861 μg/L wine, depending on MLF-container and barrel toasting. Within this family of esters, it was the only volatile present at above-threshold levels (30 μg/L) (Guth, 1997). With regard to the ethyl esters branched acids, the ethyl isobutyrate and the ethyl 3-methylbutanoate exhibited 9.6 and 10.4-fold higher concentrations than the corresponding odor thresholds, conferring hints of lemon and pineapple to the wine. Two-way ANOVA results revealed that only the MLF-container had a significant influence (p < 0.01) on the fruity volatile composition of Chardonnay wines. In the case of T2 and T3 toastings, tank-fermented wines exhibited the highest values for all esters considered (p < 0.05), apart from isobutyl acetate which was present at 22 – 31% greater concentrations in MLF-barrel wines, and isoamyl acetate and/or ethyl 3-methylbutanoate which depicted similar concentrations for both MLF-modalities (Table 2). The T1 toasting reflected the same general


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Table 2 Fruity volatile composition (n = 4) of Chardonnay wines of the two MLF modalities (in stainless steel tanks and in barrels) after 12 months of ageing in oak barrels representing different toasting processes (T1, T2, T3).

Ethyl esters of straight-chain fatty acids Ethyl butyrate 20a⁠

Odour descriptor

T1 toasting

T2 toasting

MLF in tank

MLF in barrel

462,8 ± 24,2 a

441,0 ± 0,3 a

1157,2 ± 95,8 a

1119,4 ± 4,9 a

1965,3 ± 144,5 a 1254,4 ± 71,5 a

1913,0 ± 13,4 a 1068,2 ± 13,3b

T3 toasting


Odour threshold (μg/ L)

MLF in tan

MLF in barrel

MLF in tank

MLF in barrel

450,8 ± 0,6 a

425,0 ± 0,5b

447,7 ± 4,3 a

420,3 ± 1,4b

1148,3 ± 0,7 a

1108,7 ± 0,9b

1109,3 ± 5,0b

1997,3 ± 25,3 a 1254,3 ± 4,8 a

1896,9 ± 11,3b

1162,5 ± 6,6 a 2061,3 ± 2,3 a 1285,6 ± 3,0 a

38,8 ± 0,3 a 4,2 ± 0,1 a 824,3 ± 11,1 a

50,6 ± 0,0b 3,8 ± 0,0b 814,1 ± 2,7 a

49,6 ± 0,9b 3,6 ± 0,1b 706,7 ± 0,1b

6,9 ± 0,0 a 90,9 ± 1,3 a

4,7 ± 0,0b 80,7 ± 4,6b

40,6 ± 1,2 a 4,0 ± 0,1 a 861,3 ± 10,3 a 7,9 ± 0,1 a 98,9 ± 2,7 a

Ethyl hexanoate


Ethyl octanoate


Ethyl decanoate


kiwi, strawberry, fruity green apple, fruity pinneaple, pear, floral fruity, floral

Isobutyl acetate Butyl acetate Isoamyl acetate

1600c⁠ 1880d⁠ 30a⁠

fruity fruity banana

41,6 ± 2,8 a 4,1 ± 0,0 a 843,9 ± 52,2 a

51,0 ± 0,0b 3,8 ± 0,2 a 714,5 ± 0,8 a

Hexyl acetate 2-phenylethyl acetate

1500e⁠ 250a⁠

pear, apple floral

7,1 ± 0,6 a 93,9 ± 10,4 a

3,5 ± 0,0b 67,0 ± 0,9b

lemon, kiwi, fruity apple, kiwi, fruity lemon, pinneapple, fruity

146,5 ± 8,0 a

141,9 ± 0,5 a

148,5 ± 0,7 a

142,5 ± 2,0b

147,2 ± 1,1 a

140,4 ± 0,9b

13,6 ± 0,7 a

12,4 ± 0,0 a

13,5 ± 0,3 a

12,7 ± 0,2b

13,6 ± 0,1 a

12,5 ± 0,1b

30,6 ± 1,9 a

31,4 ± 0,1 a

30,9 ± 0,5 a

31,5 ± 0,1 a

31,1 ± 0,1 a

31,4 ± 0,3 a


Higher alcohol acetates

Ethyl esters branched acids Ethyl isobutyrate 15b⁠ Ethyl 2-methylbutanoate Ethyl 3-methylbutanoate

18b⁠ 3b⁠

1030,2 ± 29,9b

1896,5 ± 17,9b 1015,1 ± 64,0b

3,3 ± 0,1b 68,6 ± 5,1b

All results were expressed in μg/L wine. T, toasting. MLF, malolactic fermentation. For each toasting method and volatile compound, means followed by different letters showed significant differences (p < 0,05) between MLF-containers. a⁠ 10% aqueous EtOH (v/v), Guth (1997). b⁠ 11% aqueous EtOH (v/v), 7 g/L glycerin, 5 g/L tartaric acid, pH adjusted to 3.4 with 1 M NaOH, Ferreira et al. (2000). c⁠ 10% aqueous EtOH (v/v) at pH 3,2, Ferreira et al. (2002). d⁠ 12% aqueous EtOH (v/v), Gómez-Míguez et al. (2007). e⁠ 10% aqueous EtOH (v/v), Étiévant (1991).

trend, even if the fruity aroma profile of both MLF-modalities did not differ significantly (p > 0.05) for most of esters. These differences in ester concentrations depending whether MLF takes place in barrels or tanks, may be explained by the esterase activity of LAB. It has been shown that bacteria performing MLF are able to alter concentration of esters that are present in wine after alcoholic fermentation, either by producing or by consuming them, changing flavor and organoleptic perception of wine in both cases (Swiegers, Bartowsky, Henschke, & Pretorius, 2005). The obtained results suggest that the particular conditions given by each MLF-container (i.e. micro-oxygenation process and release of woody compounds into wine when MLF occurred in barrels but not in tanks) may either enhance synthesis (in tanks) or hydrolysis (in barrels) of esters during this winemaking step. Thus, as observed for the woody aroma, the fruity aroma is more impacted by the MLF modality than the toasting method.

T2 barrels led to wines characterized with less aromatic intensity and perceived as less bitter, whereas T3 barrels led to wines described as the least sweet ones. In the case of MLF-tank wines, significant differences with regard to the barrel toasting were only observed for the olfactory nuts descriptor (p < 0.05). Particularly, T1- and T2-barrel wines were perceived to release the highest intensity of nuts. Barrel toasting notably conditioned judges’ preference, when tasting wines of both MLF-modalities. From an olfactory point of view, no preference was reported for T1 and T3 toastings, whereas MLF-tank modality was slightly more appreciated (55%) for T2 toasting. In mouth, wine that went through MLF in stainless steel tank was preferred in the case of T3 toasting (59%). Meanwhile, for T1 and T2 toastings, MLF-barrel wines were further more appreciated (55% and 58%, respectively). The triangle test and the bilateral paired comparison test were carried out on both barrel- and tank-fermented wines for each toasting method separately. From the triangle test, judges found significant differences (p < 0.05) between wines of both MLF-modalities for the three toasting methods considered. In the case of T2 toasting, according to the results of the bilateral paired comparison test, these differences were not significantly correlated to any of the evaluated attributes (p > 0.05). In contrast, according to the results of the bilateral paired comparison test, for the T1 and T3 toastings the differences detected between barrel- and tank-fermented wines were significantly related to the nuts flavor (higher intensity for MLF-barrel wines). This behavior of enhanced hazelnut notes when MLF is carried out in oak barrels, has been previously described for Chardonnay wines from Burgundy (Sauvageot & Vivier, 1997), and recently investigated from a

3.4. Sensory evaluation of wines

Within each MLF-modality, preliminary preference test results showed that wines from T2 barrels were generally the most appreciated (42 – 63%), whereas wines from T1 barrels were the least preferred (42 – 56%). Average values of both olfactory and gustative descriptors of Chardonnay wines after 12-months ageing in barrels (with T1, T2 and T3 toastings), are depicted in spider web diagrams at Fig. 3, separately for each MLF-modality. In the case of barrel-fermented wines, the toasting level led to significant differences (p < 0.05) with regard to the aromatic intensity, sweetness and bitterness attributes. Specifically,


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M.R. González-Centeno et al.

Fig. 3. Descriptive sensory evaluation of Chardonnay wines after 12 months of ageing in oak barrels representing different toasting processes (T1, T2, T3): olfactory attributes of both MLF-tank (A) and MLF-barrel (B) modalities, and gustatory attributes of both MLF-tank (C) and MLF-barrel (D) modalities. * means significant differences at p < 0.05.

chemical point of view in model wine (Gros et al., 2017). Specifically, Gros et al. (2017) have identified five pyrroles reminiscent of hazelnut in Chardonnay wines, whose presence might be partly due to their release from oak wood during Chardonnay storage in barrels. In the case of T1 toasting, tasters also reported significant differences (p < 0.05) between both MLF-modalities regarding the olfactory descriptors citrus fruits and white flowers, pointing out a higher intensity for MLF-tank wines. This might be attributed to the slightly higher ester concentration of the tank-fermented modality wine. In the case of T2 and T3 toastings, both MLF modalities were not differentiated with

regard to those olfactory descriptors (p > 0.05), even if tank-fermented wines also exhibited greater ester contents. 4. Conclusions

Overall, the present research reports a first examination of the chemical and sensory evolution of Chardonnay wines caused by the use of oak wood during MLF. To the best of the authors’ knowledge, no studies addressing this issue for a long ageing period (12 months) have been previously published in the literature for white wines.


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Food Chemistry xxx (2018) xxx-xxx

Barrel toasting impacts ellagitannin content to a higher extent than MLF-container. The availability and extractability of these oak wood compounds are directly dependent on barrel toasting, observing a particular ellagitannin profile for each wine considered. On the other hand, MLF-container has generally greater influence on the aroma composition of Chardonnay wines. When comparing both MLF-modalities, barrel-fermented wines presented higher amounts of whiskey-lactones and vanillin, but slightly lower concentrations of certain fruity volatiles. The barrel toasting seemed to play an important role in sensory differences between Chardonnay wines from both MLF-modalities. Depending on the style targeted for Chardonnay wines, producers may obtain interesting higher nuts aroma intensity when MLF undergoes in barrel, or greater citrus and floral aromas when wines go through MLF in stainless steel tanks. In any case, results suggested that the use of barrels as the MLF-container i) does not change the perception of the main aromas (peach, apricot and flint) defining Chardonnay character; ii) does not impact the aromatic intensity and persistence, sweetness, acidity, mouthfeel volume and/or bitterness attributes of wine; and iii) does not confer a significantly higher overall woody aroma which might mask the fruity, floral and mineral character of Chardonnay wine.


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Conflict of interest The authors declare that there is not is no conflict of interest. Acknowledgements

The authors gratefully acknowledge Tonnellerie Nadalié cooperage (Ludon-Médoc, France) for providing the oak barrels and the financial support for this research. They also thank Domaine Costa Lazaridi for supplying the wine samples, as well as the judges who participated in the sensory analyses and Dr. S. Tempère for her advice in the statistical treatment of the sensory results. References

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