Replacement of sulfur dioxide by hydroxytyrosol in white wine: Influence on both quality parameters and sensory

LWT - Food Science and Technology 65 (2016) 214e221

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Replacement of sulfur dioxide by hydroxytyrosol in white wine: Influence on both quality parameters and sensory
n c, Bele
n Puertas a, Rafaela Raposo a, María J. Ruiz-Moreno b, Teresa Garde-Cerda
M. Moreno-Rojas b, Pilar Zafrilla d, Ana Gonzalo-Diago c, Raul F. Guerrero b, Jose Emma Cantos-Villar b, *
n y Formacio
n Agraria y Pesquera (IFAPA) Rancho de la Merced, Ctra Trebujena, km 2.1, 11471 Jerez de la Frontera, Spain Instituto de Investigacio
n y Formacio
n Agraria y Pesquera (IFAPA) Alameda del Obispo, Avd Men
Instituto de Investigacio endez Pidal, 14004 Cordoba, Spain ~ o, Instituto de Ciencias de la Vid y del Vino, Gobierno de La Rioja-CSIC-Universidad de La Rioja, Carretera de Burgos Km. 6, Finca La Grajera, 26007 Logron Spain d
lica San Antonio, Campus de Los Jero
nimos, s/n Guadalupe, 30107 Murcia, Spain Universidad Cato a

b c

a r t i c l e i n f o

a b s t r a c t

Article history: Received 12 May 2015 Received in revised form 21 July 2015 Accepted 2 August 2015 Available online 5 August 2015

The feasibility of two hydroxytyrosol commercial products as an alternative to sulfur dioxide (SO2) in Sauvignon wines was evaluated. The hydroxytyrosol enriched products came from synthesis and olive waste. For this purpose wines elaborated with those products were compared with control ones elaborated with SO2. Enological parameters, color related parameters, antioxidant activity, volatile compounds, sensory analysis and olfactometric profile were determined in wines. Moreover, the evolution of wines after bottling was evaluated over six months. No significant differences were found in enological parameters and volatile composition (esters, alcohols and acids). However, significant differences were observed in color related parameters, antioxidant capacity, sensory analysis and olfactometric profile. Hydroxytyrosol-enriched wines resulted more colored and with higher antioxidant activity. Their main sensorial attributes did not correspond with the typical for Sauvignon blanc wines, which was related with a decrease in the odor intensity of the volatile thiol 3-mercaptohexyl acetate. © 2015 Elsevier Ltd. All rights reserved.

Keywords: Aroma Color Hydroxytyrosol Olfactometry Wine quality

1. Introduction The most widely preservative used in wine industry is sulfur dioxide (SO2). Its antioxidant and antimicrobial properties make it essential nowadays. SO2 has been used to inhibit polyphenol oxidase activity during winemaking, as well as to control the onset of undesirable fermentations such as acetic or malolactic fermentation (Guerrero & Cantos-Villar, 2015). However, the use of SO2 has also drawbacks. Several human health risks, including dermatitis, urticarial, angioedema, diarrhea, abdominal pain, bronchoconstriction and anaphylaxis, have been associated to SO2. It is important to reduce the amount of SO2 in wine since this compound is found in many food products as a food preservative and the amount consumed is accumulated in the organism (Vally, Misso, & Madan, 2009). Consequently, the International Organization of Vine and Wine (OIV) has been progressively reducing the

* Corresponding author. E-mail address: [email protected] (E. Cantos-Villar). http://dx.doi.org/10.1016/j.lwt.2015.08.005 0023-6438/© 2015 Elsevier Ltd. All rights reserved.

maximum SO2 recommended concentration authorized in wines,
wines and 160 mg/L which is nowadays 210 mg/L for white and rose for red wines (OIV, 2012). Moreover, SO2 addition in wine can produce organoleptic alterations in the final product, neutralize the aroma and even produce characteristic aroma defects. For above reasons the research on alternatives to SO2 has been enhanced. Some emerging technologies, also called green technologies, have been proposed as possible alternatives to SO2. Pulsed electric field, ultrasounds, high pressure and ultraviolet light have been tested in wines. However, further research is necessary to know in more detail the effect of those treatments on the sensorial properties of wines, and to validate the applicability of these technologies in wineries. Some chemical compounds have been also investigated: colloidal silver complex, dimethyl dicarbonate, ascorbic acid, hypophosphorous acid, thiodipropionic acid, Trolox C, stannous chloride, Sporix, sodium hypochlorite and even natural products such as lysozyme and bacteriocins (Santos, Nunes, Saraiva, & Coimbra, 2012). Among them, the use of phenolic compounds has been proposed as a promising alternative. For example, enological

R. Raposo et al. / LWT - Food Science and Technology 65 (2016) 214e221

tannins combined with lysozyme were added in alcoholic fermentation (Sonni, Cejudo Bastante, Chinnici, Natali, & Riponi, 2009), and rich extracts in polyphenols from almond skin and eucalyptus leave have been proved in Verdejo wines during aging
lez-Rompinelli et al., 2013). However an alternain barrels (Gonza tive which completely substitute SO2 in winemaking has not been found yet. Hydroxytyrosol (HT) is a phenylethyl alcohol which shows high antioxidant and antimicrobial capacity. HT is naturally found in wine in a wide concentration range. In white wines, values ranged
ndez-Mar, Mateos, García-Parrilla, from 1.75 to 45 mg/L (Ferna Puertas, & Cantos-Villar, 2012). HT, among other olive oil polyphenols, has been recently accepted as protective compound against oxidative damage (EFSA, 2011). In a previous study the antioxidant activity, antimicrobial activity and olfactometric profile of an olive mill waste extract with high HT concentration was evaluated (Ruiz-Moreno et al., 2015). It was concluded that the extract was a suitable source of both antioxidants and antimicrobials, although its odorants may contribute negatively to wine. Based on these results, in the current work two different HT extracts, were tested as a possible alternative to SO2 in white wine. The aim of this study was to evaluate the feasibility of hydroxytyrosol as an alternative to SO2 in white wines. Enological quality parameters, color related parameters, antioxidant capacity, volatile composition, olfactometric profile and sensory wine properties were evaluated. 2. Materials and methods 2.1. Chemicals Analytical grade methanol and formic acid were supplied by Panreac (Barcelona, Spain). Chemical standards: hydroxytyrosol, Trolox (6-hidroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid), 2,20-azobis(2-amidinopropane) dihydrochloride (AAPH), 2,2diphenyl-1-picrylhydrazyl (DPPH), K(OH) solution, dichloromethane (LiChrosolv quality), aroma standards and alkane solution (C7eC40) used for identification were purchased from SigmaeAldrich (Steinheim, Germany). Anhydrous sodium sulfate was obtained from Panreac (Barcelona, Spain). Ultrapure water from a Milli-Q system (Millipore Corp., Bedford, MA) was used throughout this research. 2.2. Hydroxytyrosol commercial products Two products based on HT were used in the present study. The first, HT product was produced by chemical and enzymatic synthesis with analytical purity (>99%) (Seprox Biotech, Spain), hereinafter called as HTB. It is generally recognized as safe (GRAS). The second one, was a natural extract from olive byproducts, whit a richness of 26% HT (Hytolive®, Genosa I þ D, Spain), hereinafter referred as HTG. 2.3. Winemaking A 600 kg of Sauvignon blanc grapes were harvested, destemmed, crushed and pressed. After pressing, enzymes were added into the must (2.5 mL/hL, Enartis ZYM Blanco L, Italy), dejuiced (24 h at 4  C) and placed in a 300 L steel vessel, which had been previously filled up with nitrogen. Alcoholic fermentation (AF) was then started after yeasting (Aroma White, Italy). AF was developed during 13 days under control temperature (18  C). CO2 was added daily to assure reductive conditions. Afterward, wine was kept cold (15  C) under nitrogen atmosphere for another 13 days until sugar concentration was under 3 g/L. Then, the wine was divided in three

215

batches, each one in triplicate. 80 mg/L of SO2 (Solfosol, SepsaEnartis, Spain) were added to CT wines, 80 mg/L of HT synthetic to HTB wines, and 308 mg/L of Hytolive (for 80 mg/L of HT) to HTG wines. This concentration was selected in agreement with antioxidant activity of hydroxytyrosol (Ruiz-Moreno et al., 2015) and previous sensory studies (data not shown). Wines were stabilized in a cold chamber during two months. Subsequently, the wines of each batch were racked, filtered (Optical XL 10W, Millipore, France) and bottled. Bottled wines were stored under control conditions (16  C, 80% RH) during 6 months. Sampling was conducted after addition of antioxidants (end of AF), at bottling and after six months of storage in bottle. 2.4. Enological parameters Density, ethanol, glycerine, dry extract, total and volatile acidity, pH, SO2, organic acids (acetic, citric, tartaric, malic, lactic and succinic acids) and metals (Na, K, Ca, Cu, Fe and Zn) were determined at bottling following the official analytical methods established by the International Organization of Vine and Wine (OIV, 2012). 2.5. Color related parameters Color intensity (D.O. 420 nm þ D.O. 520 nm) and hue (D.O. 420 nm/D.O. 520 nm) were determined by spectrophotometric measures (Lambda 25, PerkineElmer, USA). Colorimetric measurements were registered with a Konica-Minolta CM-3600d spectrophotometer (Osaka, Japan), using 20 mm path length glass cells and distilled water as reference. The CIELab parameters (L*, a*, b*) were determined by using the software SpectraMagic v.3.61G (Cyberchrome Inc, Minolta Co. Ltd), following the recommendations of the Commission Internationale de L'Eclariage (CIE): the standard observer (D10 ) and the standard illuminant (D65). Color differences (DE*ab) were calculated as the Euclidean distance between two points in the 3D space defined by L, a*, and b* (Martínez,
rez, Hita, & Negueruela, 2001). Melgosa, Pe 2.6. HPLC determination of hydroxytyrosol ~ eiro, Hydroxytyrosol was quantified as described by authors (Pin Cantos-Villar, Palma, & Puertas, 2011). Briefly, 20 mL of must or wine were analyzed by a Jasco high-performance liquid chromatographic system equipped with a diode array detector (model MD-2010), a fluorescence detector (model FP-2020), an HPLC pump module (model PU-2089), a column oven module (model CO-2060) and an auto-sampler module (AS-2050), controlled by Chrompass version 1.8 software. 2.7. Analysis of volatile compounds by gas chromatography The analysis of wines fermentative volatile compounds was
pez, Cacho, and performed by the method described by Ortega, Lo
n et al., 2014) afFerreira (2001) with modifications (Garde-Cerda ter 3 months of bottling. The extracts were injected onto a HewlettePackard (Palo Alto, CA) 6890 gas chromatograph equipped with an automatic injector and a HewlettePackard FID detector. Separation was carried out with a DB-Wax capillary column (60 m  0.32 mm I.D. x 0.5 mm film thickness; J&W Scientific, Folsom, CA). The temperature program was: 40  C for 5 min then raised up to 220  C at a rate of 3  C/min. The carrier gas was nitrogen at a flow rate of 3 mL/min. Injector temperature was 220  C and detector temperature was 280  C. Identification of compounds was carried out by comparison of their retention times with those of pure reference standards.

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fragmentation patterns with those of reference compounds or with mass spectra in the NIST 05 mass Spectral library.

2.8. Sensory analysis Descriptive sensory analysis was performed following the Sensory Profile method according to standard ISO 6564:1985 (ISO, 1985) by 11 judges. The descriptors were scored on a scale of 1e10 (1: absence of a descriptor, 10: maximum intensity). The descriptors most likely to be affected by the winemaking process were: color intensity, tonality, aroma intensity, fruity (citrus, white, tropical, stone and mature fruit), aroma defects (dirty, reduced, oxidized, yeast, chemical, rancid, acetic, acetaldehyde and ethyl acetate), savor intensity, acidity, bitter, heat, astringent, balanced and persistence. Moreover, in order to study the existence of differences among CT, HTB and HTG wines, a triangular test (ISO Standard 4120:1983) was employed (ISO, 1983). 2.9. Gas chromatography-olfactometry analyses GC analyses were carried out in wines stored 6 months in bottle using a Varian 3800 GC (Middelburg, The Netherlands) equipped with a FID and an OP275 olfactometer (GL Science Inc., Tokyo, Japan). 1 mL of extract was injected in splitless mode, being 1 min the splitless time. The columns used were DB-WAX and DB-5 from J&W Scientific, Agilent Technologies Inc. (Santa Clara, California, USA), 60 m  0.25 mm with 0.25 mm film thickness. The sample extraction and method followed has been described by authors (Sarrazin, Dubourdieu, & Darriet, 2007). The sensory panel was composed of three trained tasters, who sniffed each sample three times. Results were expressed as “modified frequency” MF (%) with the formula proposed by Dravnieks (1985). For GCeMS analysis 1 mL of extract was injected in a Trace GC Ultra gas chromatograph (Thermo Fisher Scientific S.p.A., Milan, Italy) under the same conditions described above. The detector was a mass spectrometer (ISQ single quadrupole MS, Thermo Fisher Scientific, Austin, Texas, USA) operating in EI mode (70 eV), connected to the GC with a heated transfer line at 230  C. Mass spectra were taken over the 40e350 m/z range. Thermo Xcalibur software (Thermo Fisher Scientific, San Jose, California, USA) was used for data acquisition. The odorant compounds were identified on the basis of linear retention index and a comparison of MS

2.10. Statistics Data were analyzed by one-way analysis of variance (ANOVA) on the average values. ANOVA and Least Significant Difference test (Tukey) were carried out with a significance level of p  0.05. Statistic version 9.0 (Analytical Software, Tallahassee, FL, USA) was used. 3. Results and discussion 3.1. Enological parameters In Table 1 enological parameters of wines at bottling are shown. No significant differences were found for density, ethanol, glycerine, pH, total and volatile acidity, acetic acid, citric acid, malic acid, lactic acid and succinic acids. No differences in metals (sodium, calcium, iron, copper and zinc) were found. Wines elaborated with HT (HTB and HTG) showed higher dry extract than CT wines, probably due to tartaric acid. CT wines showed small but significant lower tartaric acid and potassium than HTB and HTG wines. Regarding antioxidants, only CT wines contained free sulfur dioxide. HT was present in low amount in CT wines and around 80 mg/L in wines elaborated with HT (HTB and HTG), which meant that HT was not lost during racking and filtration. Thus, it can be concluded that minimal differences were observed among wines with regards to enological quality parameters. This is in agreement with authors ~ as, García-Romero, Huertas-Nebreda, & Go
mez(Izquierdo-Can Alonso, 2012), who studied the use of colloidal silver complex instead of SO2 in the elaboration of Merseguera white wines. The authors found small but significant differences in alcohol, total and volatile acidity, as well as in malic and citric acids. In contrast, higher differences respect to the control were found in pH and total acidity of Parellada white wines when pulsed electric fields were
n, Marselle
s-Fontanet, Ariasused to stabilize them (Garde-Cerda Gil, Ancín-Azpilicueta, & Martín-Belloso, 2008). Enological parameters permit to confirm that wine comply with

Table 1 Enological parameters of Sauvignon blanc wines at bottling.

Density relative Ethanol (%vol) Glycerine (g/L) Dry extract (g/L) Total acidity (g/L TH2) pH Volatile acidity (g/L AcH) Acetic acid (g/L) Citric acid (g/L) Tartaric acid (g/L) Malic acid (g/L) Lactic acid (g/L) Succinic acid (g/L) HT (ppm) Total SO2 (mg/L) Free SO2 (mg/L) Na (mg/L) Ca (mg/L) K (mg/L) Fe (mg/L) Cu (mg/L) Zn (mg/L)

CT

HTB

HTG

LS

0.9907 (0.0001) 12.87 (0.03) 4.95 (0.04) 15.18 (0.06)b 6.44 (0.01) 2.98 (0.03) 0.30 (0.01) 0.24 (0.01) 0.45 (0.01) 3.13 (0.06)b 2.32 (0.08) 0.07 (0.02) 0.45 (0.01) 3.92 (0.33)b 65 (1) 13 (0) <10 <50 418 (5)c <1 <0.2 <0.5

0.9908 (0.0000) 12.88 (0.03) 5.02 (0.03) 15.63 (0.09)a 6.69 (0.08) 2.95 (0.00) 0.29 (0.01) 0.23 (0.01) 0.46 (0.02) 3.42 (0.03)a 2.47 (0.06) 0.11 (0.01) 0.44 (0.01) 82.35 (0.94)a 6 (1) e <10 <50 436 (6)b <1 <0.2 <0.5

0.9908 (0.0000) 12.85 (0.01) 4.99 (0.05) 15.69 (0.12)a 6.65 (0.16) 2.96 (0.01) 0.28 (0.02) 0.24 (0.01) 0.47 (0.00) 3.43 (0.01)a 2.44 (0.07) 0.09 (0.01) 0.44 (0.00) 74.16 (1.48)a 6 (1) e <10 <50 450 (4)a <1 <0.2 <0.5

e e e ** e e e e e *** e e e *** *** *** e e *** e e e

CT, SO2 addition wine; HTB, synthetic hydroxytyrosol addition wine; HTG, hytolive addition wine; HT, hydroxytyrosol; TH2, tartaric acid; AcH, acetic acid. Standard deviation between brackets. Different superscript letters (a, b or c) for the same parameter denote significant differences (p < 0.05). Analyses of variance, level of significance (LS): * (p < 0.05), ** (p < 0.01), *** (p < 0.001).

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legality and quality standards. Twenty one enological parameters were analyzed. Only dry extract, tartaric acid and potassium where significantly different. However, they were within legal, quality and usual range in white wines. 3.2. Evolution of color related parameters Significant differences were observed in wine color (Table 2, end AF). HTB and HTG wines showed higher color intensity (CI), a* and b* than CT wines. Meanwhile hue resulted lower in HTB and HTG wines than in CT wines. Higher a* and b* values in HT wines implied wines less green and more yellow in relation to CT wines. Differences were similar to those found at “end AF” for all parameters except for L* at bottling. HT wines resulted darker (lower L*) than CT wines. Wine color parameters, especially color intensity, evolved during the storage in bottle (6 months) in all assays. a* Decreased and b* increased during the aging according with other authors (Boido, Alcalde-Eon, Carrau, Dellacassa, & Rivas-Gonzalo, 2006). In HT wines b* increased importantly whereas differences in a* among wines disappeared with storage. In order to establish whether the observed changes in the chromatic parameters were visually relevant, the color differences (DE*) between HT and CT wines were calculated. DE* values  3 CIELab units have been related to human-eye perceptible differ
mez-Míguez et al., 2007). Differences at the end ences in color (Go of AF were lower than 3; at bottling differences were lower than 3 only for HTB wines; after 6 months of storage in bottle, the differences were higher than 3 in both HTB and HTG wines (DE*ab ¼ 4.38 and 5.60 CIELab units, respectively). Other authors have described important differences in wine color when alternatives to SO2 were ~ as et al., 2012). tested (Falguera, Forns, & Ibarz, 2013; Izquierdo-Can ~ as et al. (2012) determined color parameters in Izquierdo-Can bottled wines. White wines treated with colloidal silver complex changed importantly the coordinates CIELab. Falguera et al. (2013) also described high differences when compared color parameters of irradiated wines with control ones. As far of our knowledge, it is the first time when studying alternatives to SO2 that the evolution of white wine color with storage in bottle has been described. 3.3. Volatile compounds The volatile compounds were grouped into 3 categories (Fig. 1): esters (n ¼ 12), alcohols (n ¼ 9) and fatty acids (n ¼ 6). Within esters (Fig. 1A), ethyl acetate followed by ethyl lactate were the most abundant, which values above 15 and 5 mg/L, respectively. Only ethyl lactate showed significant differences due to the added antioxidant. This ester provides milky notes to wines and is responsible for the volume sensation of wines. Nevertheless, ethyl lactate concentration in white wines is low and it was present below of its perception threshold (Campo, Ferreira, Escudero, &

217

Cacho, 2005). Acetate ester of higher alcohols, mainly isoamyl acetate and 2-phenylethyl acetate, and ethyl esters of fatty acids, mainly ethyl hexanoate and ethyl octanoate were not affected by the type of antioxidant used during the storage in bottle. These compounds are considered important contributors to young wine aroma and exhibit floral and fruity odors. Isoamyl acetate was found in the wine at concentrations close to its perception threshold (1 mg/L) () and 2-phenylethyl acetate at concentrations above its threshold level (0.25 mg/L). Regarding ethyl esters of fatty acids, the concentration of ethyl hexanoate was above its perception threshold (0.08 mg/L) while the concentration of ethyl octanoate was below of its perception threshold (0.58 mg/L) (Peinado, Moreno, Bueno, Moreno, & Mauricio, 2004). The alcohols (Fig. 1B) showed significant differences due to the antioxidant only in methionol, isoamyl alcohols, and 2phenylethanol. Control wines showed lower concentration of methionol than HTB and HTG wines, which did not show significant differences between them. Methionol is synthesized by yeast during fermentation and it is known to be very sensitive to oxidation (Dombre, Rigou, Wirth, & Chalier, 2015). Thus, methionol was more oxidized in absence of SO2. Methionol exhibits boiled potato notes and its concentration was under the odor threshold in all wines
pez, & Cacho, 2002). In the (1 mg/L) (Ferreira, Ortín, Escudero, Lo case of isoamyl alcohols, control wines had higher concentration of these compounds than HT wines, once again, with no differences between them. The concentration of 2-phenylethanol was higher in control than in HTG wines, without showing significant differences with HTB wines. Total alcohol content was significant lower in wines without SO2. This result agrees with Garde-Cerd
an and Ancín-Azpilicueta (2007) and Sonni et al. (2009) who found that the suppression of SO2 caused a decrease on the total alcohol content in wines. Alcohols have intense odors that play a role in wine aromas. At low concentrations (less than 300 mg/L), they contribute to the wine aromatic complexity. At high levels, their penetrating odors mask the wine aromatic finesse. The concentration of total alcohols in wines was below to 300 mg/L, so these compounds probably enhanced the wine aroma. Few differences were found for fatty acids (Fig. 1C). Only butyric acid was affected by the added antioxidant. Its concentration was lower in HTG wines than in CT and HTB wines. Other authors did not find differences in the concentration of fatty acids after 3 months of white wines aging in bottle with and without SO2
n & Ancín-Azpilicueta, 2007). Fatty acids contribute to (Garde-Cerda the fresh flavor of wine, but, in excess, (>20 mg/L), became an
n et al., 2005). unpleasant flavor, impairing wine aroma (Pozo-Bayo They also help to modify the perception of other taste sensations. Minor differences in wine volatile composition have been also described by authors who tested other alternatives to SO2 such as pulsed electric fields (Garde-Cerd
an et al., 2008), plant phenolic
lez-Rompinelli et al., 2013) and colloidal silver extract (Gonza

Table 2 Color related parameters in Sauvignon blanc wines. End AF

Bottling

CT CI Hue L* a* b* DE*

HTB c

0.044 6.299a 98.06 1.35b 6.26b

HTG b

0.059 3.811b 97.42 0.79a 8.52a 2.28

LS a

0.063 3.833b 97.22 0.80a 9.00a 2.81

*** *** e *** ***

CT

6 Months of storage HTB

c

0.032 5.344a 99.11a 1.30b 5.25c

HTG b

0.067 4.803b 98.38b 1.09a 7.53b 2.38

LS a

0.072 4.593b 98.03b 1.06a 8.08a 3.02

*** *** *** *** *** e

CT

HTB b

0.060 5.17a 98.53a 1.26 7.10b

HTG a

0.099 4.329b 97.14b 1.29 11.25a 4.38

LS a

0.110 4.309b 96.56b 1.26 12.34a 5.6

*** ** *** e *** e

CT, SO2 addition wine; HTB, synthetic hydroxytyrosol addition wine; HTG, hytolive addition wine. AF, alcoholic fermentation. CI, color intensity. DЕab*, color differences. Different superscript letters (a, b or c) for the same parameter denote significant differences (p < 0.05). Analyses of variance, levels of significance (LS): * (p < 0.05), ** (p < 0.01), (p < 0.001).

***

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Fig. 1. Volatile compounds determined in Sauvignon blanc wines, grouped according to similarity in structure: A) esters, B) alcohols, C) acids. CT, synthetic hydroxytyrosol addition wine; HTG, hytolive addition wine.

SO2 addition wine; HTB,

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(Garde-Cerd
an et al., 2014). In contrast, volatile composition of Sauvignon blanc wine (at the end of AF and after 1 year of storage) was significantly modified when SO2 was replacement by lysozyme and enological tannins (Sonni, Chinnici, Natali, & Riponi, 2011). In the present study, although AF was common for all wines, their volatile composition could have been evolved differently since the different preservatives were added at the end of AF. The results showed that the use of HT, regardless of their origin, as alternative to SO2 did not modify the wine aroma profile, and preserved volatile compounds to the same extents as SO2. 3.4. Sensory analysis Sensory analysis was conducted at bottling and after 6 months of storage. Wines at bottling (Fig. 2A) showed sensory significant differences. The highest differences were found in visual appearance parameters. Wines without SO2 (HTB and HTG wines) were described as wines with greater color intensity and hue, whereas CT wines showed a paler color. Indeed, color evolution is one of most difficult parameters to control when alternatives to SO2 in white wines are used (Guerrero & Cantos-Villar, 2015; Izquierdo~ as et al., 2012). It is worth to mention that instead DE* between Can CT and HTB wines at bottling resulted lower than 3 CIELab units, and thus not easily detected by human eye, tasters were able to detect color differences. HT wines, especially HTB wines, presented more scents of fruit (white, tropical and stone fruits). It has been described that SO2 may neutralize aromas (Guerrero & Cantos-Villar, 2015). In the present work, wines did not present any sensorial aromatic defects (dirty, reduced, oxidized, yeast, chemical, rancid, acetic, acetaldehyde or ethyl acetate). CT wines had higher savor intensity and were less balanced due to the bitter character (Fig. 2A). Sensory results of SO2 alternative application in wine have been described at bottling on the bibliography. Lysozyme-treated Riesling wine showed a greater fruity aroma intensity than the untreated control wine (Bartowsky, Costello, Villa, & Henschke, 2004), whereas wines treated with colloidal silver complex showed a loss of varietal ~ as et al., 2012). aromas (Izquierdo-Can After 6 months of bottling (Fig. 2B), differences among wines were found in visual appearance. HT wines showed higher color intensity, as described in bottling. Aromas were more evolved in HTG wines, which showed lower citrus, white and tropical fruits, and higher mature notes, in comparison with both HTB and CT wines. CT wines were scored with the highest aroma intensity. HT

219

wines became more astringent, without significant differences among both HT assays. Additionally, HTG wines were more bitter and less balanced. In summary, HT wines at bottling (especially HTB ones) showed higher scores than CT wines, but for visual appearance. After 6 months of bottling HT wines were scored lower, especially HTG. No references have been found on the sensorial effect of alternatives to SO2 on white winemaking with bottle aging. It has been recently described that wines with medium content of SO2 showed better sensory profile that these with low content of SO2, after six months of bottle storage (Panero, Motta, Petrozziello, Guaita, & Bosso, 2015). A triangular test to evaluate the differences among CT, HTB and HTG bottled wines was conducted in dark wine glasses to avoid judgments being influenced by wine color. The results showed that differences between three assays could be observed (95% significance level) in olfactory phase. An olfactometric assay was carried out to advance in the knowledge of the aromas responsible of these differences, which could not be detected by the volatile compounds analysis described above. 3.5. Olfactometric profile Olfactometric characterization obtained for each wine (CT, HTB and HTG) was listed in supplementary data (Table S1). Overall, more than 60 odoriferous perceptions were detected. To simplify, odorants not reaching a maximum GCeO scores (MF) of 30% were
et al., 2013). Odorants eliminated and considered as noise (Cullere showing maximum differences on olfactometric scores (minimum 40% MF) respect to the CT were highlighted in bold (Table S1). The analyses of variance (ANOVA) performed on olfactometric data allowed to determinate thirteen different odoriferous zones (OZs) that showed significant differences among the wine samples. Discriminant odorants were classified into three groups according to their average MF(%) values (Table 3). In the first group (MF scores > 70%), only 2-furfurylthiol (coffee) was detected, being its MF higher in CT wines than in HT ones. The second group (70 > MF(%) > 50) was composed for 8 compounds, 6 of them were identified. 4-Mercapto-4-methyl-2-pentanone and 3mercaptohexyl acetate (considered as markers of Sauvignon blanc wines) were significant lower in both HTB and HTG wines in comparison with CT ones. Linalool (floral, Muscat) and (E,Z)-nona2,6-dienal (cucumber) were also decreased in HT wines. By contrast, abhexone (curry) and an unknown compound with LRI

Fig. 2. Cobweb diagram of the sensory score for Sauvignon blanc wines at bottling (A) and after 6 months of storage (B). CT, SO2 addition wine; HTB, synthetic hydroxytyrosol addition wine; HTG, hytolive addition wine. Descriptors with * denote significant differences (p < 0.05). Analyses of variance, levels of significance: * (p < 0.05), ** (p < 0.01), *** (p < 0.001).

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Table 3 Odoriferous zones found in Sauvignon blanc wines after 6 months of storage in bottle. Gas chromatography retention data, identity, odorant description, modified frequency (%) and differences in modified frequency (%) between HT and CT wines. LRIa DB-WAX

Identity

Odorant description

DB-5

MF(%) > 70 1438 910 70 > MF(%) > 50 1363 940 1432 1137 1551 1100 1596 1156 1731 1252 1964 2261 1199 2533 1401 50 > MF(%) > 30 1611 1667 1047 1984 1491 2300 1732

2-furfurylthiolb 4-mercapto-4-Methyl-2-pentanone ethyl ciclohexanoateb Linaloolb (E,Z)- nona-2,6-dienalb 3-mercaptohexyl acetateb n.i Abhexoneb n.i n.i Phenylacetaldehydeb n.i n.i

b

DMF(%)

MF(%)

LS

CT

HTB

HTG

HTB-CT

HTG-CT

Coffee

71

41

55

30

16

**

box tree Fruity Floral, muscat Cucumber box tree Chamomile, sweet Curry Leather

68 0 57 58 62 14 34 0

50 0 36 7 37 34 50 0

55 60 43 47 0 58 38 51

18 0 21 ¡51 ¡25 20 16 0

13 60 14 ¡11 ¡62 44 4 51

*** *** ** * * * * *

Toasty, burnt Honey Fish Black pepper

0 20 7 14

41 38 0 0

14 37 38 34

41 18 7 14

14 17 31 20

* * * *

CT, SO2 addition wine; HTB, synthetic hydroxytyrosol addition wine; HTG, hytolive addition wine. In bold odorants with maximum differences on olfactometric scores n.i, not identified compound; MF, modified frequency; DMF, difference in modified frequency between HT and CT wines. Analyses of variance, levels of significance: * (p < 0.05), ** (p < 0.01), *** (p < 0.001). a Linear retention index calculated on both DB-WAX and DB-5capillary columns. b Identification based on coincidence of chromatographic retention data on capillary columns (DB-WAX and DB5) and on the similarity of odor previously reported.

1964 (chamomile, sweet) increased their intensity in HT wines. Ethyl ciclohexanoate, and an unknown compound with LRIDB-Wax 2533 (leather), were only detected in HTG wines, which made them markers for wines elaborated with hytolive. Finally, in the last odorant group (50 > MF(%) > 30), phenylacetaldehyde and an odorant zone with LRIDB-Wax 1611 (toasty) increased in HT wines. Two odorant zones with LRIDB-Wax 1984 (fish) and LRIDB-Wax 2300 (black pepper) resulted characteristic of

rez et al. (2012), odorants HTG wines. According to Alvarez-P e presenting large differences of MF(%) values (DMF > 40%) are those that have a more acute role in the perception of aromatic differences among samples. In agreement with triangular test, the olfactometric profile resulted different for CT, HTB and HTG wines. The addition of HT to wines modified its olfactometric profile, especially in HTG wines. Olfactometric profile of CT wines was characterized by the presence of the volatile thiol 3-mercaptohexyl acetate, which is responsible for the box tree aroma found in Sauvignon blanc wines (Coetzee & du Toit, 2012). HTB wines were marked by a lower intensity of (E,Z)nona-2,6-dienal and higher of LRIDB-Wax 1611 (toasty), meanwhile HTG wines were characterized by a higher intensity of LRIDB-Wax 1964 (chamomile, sweet) and the presence of ethyl ciclohexanoate (fruity) and LRIDB-Wax 2533 (leather). Thus, it seems clear that, the addition of hydroxytyrosol products decreased the varietal character of Sauvignon blanc wines, introducing odorants which might to contribute to the differences found in the sensory analysis. 4. Conclusions The results indicate that the use of hydroxytyrosol in white wine elaboration without SO2 hardly modified the enological parameters and the volatile composition of wines. However color-related parameters, sensory analysis and olfactometry analysis were modified. HT wines had higher intensity color as well as higher total score at bottling (especially HTB) but lower score after 6 months of storage in bottle (especially HTG), together with different olfactometric profile. The addition of HT commercial products decreased the varietal character of Sauvignon blanc wines. The key point of the process seems to be the evolution during the storage in bottle. The combination of those commercial products enriched in

hydroxytyrosol during winemaking with some other antioxidant additive (ascorbic acid, low SO2 concentration, glutation, tannins, etc.) at bottling should be investigated. Either suppressing or reducing the amount of SO2 is a challenge for wine industry. If it could be done by a compound naturally found in wine as hydroxytyrosol, which has recently accepted as protective compound against oxidative damage, it would achieve a healthier wine with added-value. Acknowledgments Authors thank the INIA and FEDER for their financial support (Project RTA2011-00002). Ruiz-Moreno and Guerrero thank to the European Social Fund 2007e2013 “Andalucía se mueve con Europa” for the financial support of their contracts. Appendix A. Supplementary data Supplementary data related to this article can be found at http:// dx.doi.org/10.1016/j.lwt.2015.08.005. References

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