Gamma irradiation as pre-fermentative method for improving wine quality

Accepted Manuscript Gamma irradiation as pre-fermentative method for improving wine quality Marin Mihaljević Žulj, Luna Maslov Bandić, Ivana Tartaro Bujak, Ivana Puhelek, Ana Jeromel, Branka Mihaljević PII:

S0023-6438(18)30967-8

DOI:

https://doi.org/10.1016/j.lwt.2018.11.016

Reference:

YFSTL 7580

To appear in:

LWT - Food Science and Technology

Received Date: 16 April 2018 Revised Date:

30 October 2018

Accepted Date: 5 November 2018

Please cite this article as: Mihaljević Žulj, M., Bandić, L.M., Bujak, I.T., Puhelek, I., Jeromel, A., Mihaljević, B., Gamma irradiation as pre-fermentative method for improving wine quality, LWT - Food Science and Technology (2018), doi: https://doi.org/10.1016/j.lwt.2018.11.016. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

ACCEPTED MANUSCRIPT Gamma irradiation as pre-fermentative method for improving wine quality Marin Mihaljević Žulja,*, Luna Maslov Bandićb, Ivana Tartaro Bujakc, Ivana Puheleka, Ana Jeromela, Branka Mihaljevićc [email protected],

[email protected],

[email protected], [email protected] a

Department of Viticulture and Enology, University of Zagreb, Faculty of Agriculture,

Division of Materials Chemistry, Ruđer Bošković Institute, Bijenička 54, 10 000 Zagreb,

Corresponding Author

TE D

Croatia.

*

M AN U

Department of Chemistry, University of Zagreb, Faculty of Agriculture, Svetošimunska

cesta 25, 10 000 Zagreb, Croatia. c

SC

Svetošimunska cesta 25, 10 000 Zagreb, Croatia. b

[email protected],

RI PT

[email protected],

EP

Phone: +385 01 239 3771

AC C

e-mail: [email protected]

1

ACCEPTED MANUSCRIPT Abstract

2

Merlot and Traminer (Vitis vinifera L.) grapes were subjected to gamma irradiation at the

3

panoramic 60Co source at four doses (670, 1300, 2000, 2700 Gy) and wines were produced

4

from the irradiated grapes. HPLC analysis of musts have shown a negative impact of

5

irradiation on the amino acids content. However, Merlot wines produced from irradiated

6

grapes demonstrated better extraction of the coloring matter. The concentrations of

7

anthocyanins increased with the increasing absorbed irradiation dose, while flavonols and

8

flavanols were not affected by irradiation. Irradiation with doses up to 2000 Gy increased

9

concentrations of fruity-floral aroma compounds, especially monoterpens and C13

10

norisoprenoids in wines, while a maximal dose of 2700 Gy expressed more the toasty and

11

caramel notes due to higher concentrations of furfural and furfuryl alcohols. Results

12

obtained suggest that ionizing irradiation might be a suitable method for grape treatment

13

since better chemical properties in wine could be achieved.

14

Keywords

15

Gamma irradiation; wine; amino acids; phenols; aroma

18 19

SC

M AN U

TE D

EP

17

AC C

16

RI PT

1

20 21 22

2

ACCEPTED MANUSCRIPT 1. Introduction

24

The aroma of wine is one of the most important factors that influence its organoleptic

25

characteristics as well as consumer preference (Gupta, Padole, Variyar & Sharma, 2015).

26

Some of the principal volatile compounds related to grapevine metabolism (monoterpenes,

27

C13 norisoprenoids, C6 alcohols and benzene compounds) can be found in free and bound

28

forms where free forms influence the aroma and flavor, while bound forms are mainly in

29

glycoside forms which are odorless. Another important group of compounds influencing

30

organoleptic properties of wine are the phenolic compounds: anthocyanins, tannins and their

31

polymers. The phenolic compounds are mainly extracted from grapes during maceration

32

(González-Neves, Favre, Gil, Ferrer & Charamelo, 2015; Garrido & Borges, 2013).

33

Phenolic composition of wine depends on the cultivar, viticulture practices, vinification

34

conditions and type of maceration (Garrido & Borges, 2013; Sacchi, Bisson & Adams,

35

2005). Moreover, it has also been reported that application of pulsed electric field treatment

36

resulted in increased extraction of polyphenols and anthocyanins due to better grape skin

37

permeability (Delsart et al., 2012; Puértolas, Hernández-Orte, Sladaña, Álvarez & Raso,

38

2010).

39

Gamma irradiation is a well-established noncontact physical method for food microbial

40

decontamination, preservation and shelf-life enhancement (Rodríguez-Pérez, Quirantes-

41

Piné, Contreras, Uberos, Fernández-Gutiérrez & Segura-Carretero, 2015). Radiation

42

treatment at doses of 2-7 kGy depending on condition of irradiation and the food, can

43

effectively eliminate potentially pathogenic bacteria and spoilage microorganisms without

44

compromising the safety, nutritional properties and sensory quality of the food (Farkas,

45

1998; WHO, 1997). Since this process is non-thermal, irradiation is also known as a cold-

46

pasteurisation method (Gupta et al., 2015; Alothman, Bhat & Karim, 2009). Moreover,

47

increased total phenolic compounds and antioxidant activity were observed for gamma

AC C

EP

TE D

M AN U

SC

RI PT

23

3

ACCEPTED MANUSCRIPT irradiation of different plant materials like soybeans, mushrooms, carrot and kale juice, fresh

49

cut vegetables and almond skin (Variyar, Limaye & Sharma, 2004; Huang & Mau, 2006;

50

Song, Kim, Jo, Lee, Kim & Byun, 2006; Fan, 2005; Harrison & Were, 2007). Caldwell and

51

Spayed (1989) reported that gamma irradiation of red wines has increased the color intensity

52

of the Cabernet Sauvignon wines without making any differences in their sensory quality.

53

Higher values of anthocyanins were also achieved in fresh grape pomace at 6 kGy (Ayed,

54

Yu & Lacroix, 1999, 2000). Irradiation of food products causes minimal modifications in

55

flavor, color, nutrients and taste although the levels of modification might vary depending of

56

the material composition used, irradiation dose and radiation source (Alothman et al., 2009).

57

Results by Chang (2003) showed improvement in rice wine taste quality while gamma

58

irradiation caused breakdown of aroma glycosides and enhanced volatile content in nutmeg

59

(Ananthakumar, Variyar & Sharma, 2006).

60

Currently, there is a lack of information about the influence of gamma irradiation on wine

61

quality and amino acids content in grape must. Thus, the aim of the present study was to

62

investigate the effect of gamma irradiation on the Merlot and Traminer grapes amino acids

63

composition as well as aroma profile of Traminer wines and aroma profile and polyphenol

64

composition of Merlot wines.

65

2. Materials and Methods

66

2.1 Chemicals

67

Hydrochloric acid, sodium hydroxide, boric acid and ethanol were of chemical purity and

68

obtained from Kemika (Zagreb, Croatia). Methanol, dichloromethane, acetonitrile and

69

dimethyl sulfoxide were obtained from J.T. Baker (Derventer, Netherlands). All other

70

chemicals including standards of amino acids, polyphenols and aroma were obtained from

71

Sigma-Aldrich (Steinheim, Germany) in the highest commercially available grade of purity.

AC C

EP

TE D

M AN U

SC

RI PT

48

4

ACCEPTED MANUSCRIPT 72

2.2 Materials

74

The grapes of Merlot and Traminer (Vitis vinifera L.) variety were manually harvested at the

75

experimental station of Faculty of Agriculture in Zagreb. Immediately after harvest the

76

grapes were packed (600 g) in cardboard boxes and transported for the gamma radiation

77

treatment.

78

2.3 Gamma irradiation of grapes

79

Grapes in boxes were gamma irradiated at the panoramic

80

Chemistry and Dosimetry Laboratory at the Ruđer Bošković Institute (Zagreb, Croatia). The

81

dose rate and the absorbed dose were established using an ethanol-chlorobenzene (ECB)

82

dosimetry system (40 %, v/v chlorobenzene in ethanol) (Ražem, Anđelić & Dvornik, 1985).

83

After irradiation of grapes at estimated dose rate of 6.4 Gy/min the irradiation doses applied

84

were measured to be 670, 1300, 2000 and 2700 Gy. Irradiation was performed under

85

ambient atmosphere and chamber temperature of 18°C.

86

2.4 Vinification procedure

87

After gamma irradiation grapes underwent the vinification procedure separately depending

88

on their variety. The Traminer grape juice obtained by pressing after de-stemming and

89

crushing was stored in glass bottles for settling, while Merlot pomace was stored in plastic

90

containers. Free SO2 was adjusted to 50 mg/L in both cases using a 5% solution of sulfurous

91

acid. After 24 h clear Traminer grape juice was separated from sediment and inoculated

92

with Saccharomyces bayanus yeast strain EC1118 (Lallemand, Canada) at the level of

93

5×106 cells/mL as well as Merlot pomace. Fermentations were carried out in controlled

94

environment at 15 °C. The cap formed at Merlot pomace during maceration was punched

95

daily and pomace was pressed after five days. After completion of fermentation samples

SC

RI PT

73

Co source in the Radiation

AC C

EP

TE D

M AN U

60

5

ACCEPTED MANUSCRIPT were racked from lees and sulfurized. Free SO2 in young wines was adjusted to 50 mg/L and

97

the wines were stored at 15 °C in a wine cellar. All treatments were carried out in triplicate.

98

2.5 Chemical analysis

99

Basic physicochemical parameters were analyzed in wines (alcohol, sugar free extract,

100

reducing sugars, total acidity, volatile acidity and pH) using methods proposed by the

101

International Organization of Vine and Wine (O.I.V., 2007).

102

2.5.1 Amino acid analysis

103

2.5.1.1 Preparation of standard solution, reagents, and sample derivatization

104

A method for determination of amino acids was applied according to Pripis-Nicolau, de

105

Revel, Marchand, Anocibar Beloqui & Bertrand (2001), modified for our analysis. Standard

106

solutions of amino acids were prepared in purified water. Few drops of 1 M HCl were added

107

to dissolve amino acids to prepare a stock solution. Mixtures of standard solutions for

108

calibration were prepared in the range from 0.50 mg/L to 100 mg/L. Calibration curves were

109

made in 5 points. Borate buffer (100 mM) was prepared and the pH 9.5 of boric acid

110

solution was adjusted with 4 M NaOH. The o-phtaldehyde thiol reagent for derivatization of

111

amino acids was prepared by dissolving 750 mg of o-phtaldehyde in 5 mL methanol, and

112

0.5 mL of 2-sulfanylethanol was added. The solution was made up to 50 mL with borate

113

buffer. The iodoacetic solution of 70 mg/L of iodoacetic acid in borate buffer was prepared

114

and pH was adjusted to 9.5 with 4 M NaOH.

115

2.5.1.2 HPLC analysis

116

HPLC analysis were performed using Agilent 1100 Liquid Chromatograph, equipped with a

117

fluorescence detector (Agilent 1200). The excitation and emission wavelengths were 356

118

nm and 445 nm, respectively. Separation of amino acid derivatives was obtained by

AC C

EP

TE D

M AN U

SC

RI PT

96

6

ACCEPTED MANUSCRIPT Lichroshere RP 18 column (125 mm × 4mm × 5 μm). Precolumn derivatization was done

120

using o-phtaldialdehyde and iodoacetic acid. Mobile phase A was a mixture of 23 % of 250

121

mM disodium hydrogen phosphate, 20 % of 250 mM propionic acid and 2 % of dimethyl

122

sulphoxide adjusted to pH 6.65 with 4 M NaOH followed by the addition of 6.5 % of

123

acetonitrile and 48.5 % of water. Mobile phase B was a mixture of 40 % of acetonitrile, 33

124

% of methanol, 7 % of dimethyl sulphoxide and 20 % of water. Flow rate was 0.8 mL/min.

125

Run time was 125 min. A sample of must was filtered through 0.45 μm PTFE (Phenomenex,

126

USA) membrane filters prior to analysis.

127

2.5.2 Aroma analysis

128

2.5.2.1 Sample preparation for GC/MS analysis

129

The extraction of aroma compounds was carried out according to Lopez, Aznar, Cacho &

130

Ferreira (2002). Cartridges containing 200 mg LiChrolut EN sorbent were preconditioned

131

with 4 mL of dichloromethane, 4 mL of methanol, and 4 mL of ethanol: water mixture (13.5

132

%, v/v). Fifty milliliters of wine were passed through the SPE cartridge and dried in vacuum.

133

The analytes were recovered by elution with 800 µL of dichloromethane. Ten microliters of

134

internal standard (50 mg/L, 2-octanol) were added over the eluted sample.

135

2.5.2.2 GC/MS analysis

136

The prepared extract was injected to an Agilent 6890 gas chromatograph coupled with 5973

137

mass selective detector, equipped with an Agilent 6890 autosampler. The GC column was

138

ZB-WAX from Phenomenex, Torrance, USA, 60 m × 0.32 mm i.d., with 0.50 µm film

139

thickness. The carrier gas was helium (Messer, Zaprešić, Croatia), 5.5 grade at a constant

140

flow rate of 1 mL/min. The injection volume was 2 µL in splitless mode. The injector

141

temperature was 250 °C. The column temperature program was as follows: initial hold for 5

142

minutes at 40 °C, followed by a 2°C/min to 240°C, and then kept for 20 minutes. The

AC C

EP

TE D

M AN U

SC

RI PT

119

7

ACCEPTED MANUSCRIPT temperature of the transfer line was 230 °C. The temperatures of the ion source and

144

quadrupole were 230 °C and 150 °C, respectively. The mass spectrometer was operated in

145

electron ionization mode at 70 eV with selected ion monitoring (SIM). Identification was

146

performed by comparing retention times and mass spectra with those of pure standards and

147

with mass spectra from NIST05 library. Linear retention indices (relative to n-alkanes) were

148

calculated and compared to those from literature. Standard five-point calibration curves

149

(based on quantification ions) were constructed.

150

2.5.3 Polyphenols analysis

151

2.5.3.1 Sample preparation

152

The wines (1 mL) were filtered through a 0.45 µm PTFE membrane syringe filter (Phenex,

153

Phenomenex, USA), and the samples were injected in triplicates for HPLC analysis.

154

2.5.3.2 HPLC analysis

155

Detection and quantitative analysis of phenolic compounds were carried out using an

156

HPLC-DAD/FLD method as described by (Tomaz & Maslov, 2016). A HPLC model 1100

157

(Agilent Technologies, USA) equipped with binary pump, an auto sampler, diode array

158

detector and Agilent 1200 fluorescence detector was used. Mobile phases consisted of (A)

159

water/phosphoric acid

160

(50/49.5/0.5, v/v/v). Separation of phenolic compounds was carried out using a Luna

161

Phenyl-Hexyl (Phenomenex, USA) column (250 mm x 4.6 mm i.d., 5 μm particle size) with

162

a Phenyl guard column (4.0 x 3.0). The column was thermostated at 50 °C. The injection

163

volume was 20 μL. Chromatograms and spectra were elaborated with a Chemstation data

164

system (Agilent Technologies, USA). Identification of compounds was based on retention

165

times, UV/Vis spectra and fluorescence using external standards. Stock solution of each

166

polyphenol standard was prepared by weighing and dissolving in methanol. Mixtures of

EP

TE D

M AN U

SC

RI PT

143

v/v) and

(B) acetonitrile/water/phosphoric acid

AC C

(99.5/0.5,

8

ACCEPTED MANUSCRIPT standard solutions for calibration were prepared by diluting stock solutions in synthetic wine

168

(5 g/L of tartaric acid in water/ethanol (88:12, v/v) solution adjusted to pH 3.2 using 1 M

169

NaOH and 1 M HCl). Calibration curves were made of 5 points. Compounds were detected

170

and quantified at 518 nm for anthocyanins, 360 nm for flavonols, 320 nm for

171

hydroxycinnamic acids, 280 nm for hydroxybenzoic acids by diode array detector and at

172

excitation wavelength 225 nm and emission wavelengths at 320 nm for flavan-3-ols.

173

2.6 Statistical analysis

174

The analysis of variance (one-way ANOVA) was applied to the experimental data. Results

175

were considered significantly different if the associated p value was below 0.05. Tukey’s

176

test was applied for mean comparisons. A principal component analysis was applied to the

177

data. All statistical analyses were performed using the SAS 9.3 software (SAS Inc., Cary,

178

USA).

179

3. Results and Discussion

180

3.1 Effect of radiation on basic chemical composition of wines

181

Even though gamma irradiation showed no effect on the basic wine chemical components

182

(ethanol, pH, titratable acidity) some statistically significant (p<0.05) effect was observed in

183

volatile acidity and sugar free extract concentrations (Table 1). Merlot and Traminer wines

184

produced from grapes treated by the irradiation dose of 2700 Gy had the lowest volatile

185

acidity and among the highest concentrations of the sugar free extract. Similar results with

186

no influence of irradiation processing on ethanol concentration, pH and reducing sugars in

187

wines was observed by Gupta et al. (2015). Our results pointed out the positive influence of

188

gamma irradiation on basic wine chemical composition.

189

3.2 Effect of radiation on amino acids content in must

AC C

EP

TE D

M AN U

SC

RI PT

167

9

ACCEPTED MANUSCRIPT Amino acids have a huge impact on must fermentability and wine volatile profile since they

191

are of paramount importance for the yeasts metabolism in alcoholic fermentation and for

192

lactic bacteria in malolactic fermentation. Their concentrations can vary according to grape

193

variety and viticultural practices. From the results presented in Table 2 and 3 it can be seen

194

that Traminer musts compared to Merlot musts had higher concentrations of almost all

195

amino acids, especially arginine, glutamate and glycine which can contribute to wine aroma

196

since they can be precursors of several volatile compounds (Moreno-Arribas and Polo,

197

2009). Regardless to grape variety, in the present study gamma irradiation had a negative

198

impact on free amino acid concentrations in musts. The irradiation process reduced total

199

amino acid content in both grape varieties (Figure 1). In Traminer musts the lowest

200

concentration of 339.07 mg/L of total amino acids was obtained from grapes irradiated with

201

the dose of 1300 Gy, presenting 60 % of loss in the total amino acid concentration. No

202

significant difference was noticed among irradiation treatments at the irradiation doses of

203

670, 2000 and 2700 Gy, Unlike Traminer, in Merlot musts the difference between non-

204

irradiated samples and irradiated samples was not significant. Minimal concentration of total

205

amino acids (265.65 mg/L) measured in must was obtained from grapes irradiated with the

206

dose of 2000 Gy at which 38 % of the total amino acid concentration was reduced.

207

Considering individual amino acids, the levels of arginine and glutamate as the main sources

208

of amino acids for yeast metabolism during wine fermentation (Zoecklein, Fugelsang,

209

Gump & Nury, 1999) were strongly reduced after gamma irradiation treatment, especially

210

glutamate (47 %). Irradiation doses of 1300 and 2000 Gy had caused the strongest reduction

211

of these amino acids depending on grape variety. In Traminer musts the reduction of

212

glutamate was 58 % and 70 % for arginine at the irradiation dose of 1300 Gy, whereas in

213

Merlot musts the reduction amount for glutamate was 38 % and 54 % for arginine at the

214

dose of 2000 Gy.

AC C

EP

TE D

M AN U

SC

RI PT

190

10

ACCEPTED MANUSCRIPT Nitrogen deficiencies in grape musts can result in sluggish or stuck fermentation which in

216

our case did not happen as all sugar was fermented (Table 1). According to Kunkee (1991)

217

minimum amino acid concentration required for completion of fermentation is 140 mg/L.

218

Therefore, though reduction of amino acids was determined using irradiation at several

219

irradiation doses, the final amino acids concentrations were not bellow the needful amount.

220

There is a lack of information from literature on effect of ionizing radiation of grapes on the

221

amino acid content. Ionizing radiation can cause chemical change or destruction of free

222

amino acids in presence of water. The type and magnitude of chemical reactions observed

223

during irradiation depend on a large number of parameters such as their structure and

224

presence of water. Amino acids in proteins are also susceptible to irradiation, but reports

225

vary depending on the nature and extent of the damage. The discrepancies in literature may

226

result from variations in the experimental conditions of irradiation, such as temperature,

227

oxygen partial pressure, water content, dose and dose rate (Nadeem Chandry & Evans,

228

1971). The changes in amino acids concentration induced by irradiation are due to free

229

radicals primarily formed from the radiolysis of water (mainly the solvated electrons and the

SC

M AN U

●

TE D

230

RI PT

215

OH radicals), in association with splitting of the covalent bonds, deamination and

decarboxylation reactions of amino acids followed by chains of chemical radical reactions

232

forming other new radicals (Spinks & Woods, 1990; Bamidele & Akanbi, 2015). It is

233

evident that amino acids loss during irradiationis at a lesser extent in Merlot grapes then in

234

the Traminer grapes. This result strongly indicates the suppresion/inhibition of radical

235

reactions through stronger antioxidative potential of Merlot grapes.

236

However, previous study on edible seeds and gamma irradiation processing on amino acids

237

content showed that irradiation doses from 0 to 6000 Gy caused increase in the free amino

238

acid content (Maity et al., 2009) what is contrary to our results. In the present study, the

239

observed decrease in free amino acid content after exposure to ionizing radiation is in

AC C

EP

231

11

ACCEPTED MANUSCRIPT agreement with findings reported by Abu, Muller, Duodu & Minnaar (2005). All of these

241

observations and inconsistency between reported results together with the knowledge about

242

precise effect of ionising radiation on free amino acid content can be due to the sensitivity of

243

the exposed system to irradiation, the type of particular functional plant tissue and other

244

conditions (Maity et al. (2009).

245

3.3 Effect of radiation on phenolic compounds of Merlot wines

246

Impact of irradiation on the phenolic compounds of Merlot wines are shown in Table 4.

247

Gamma irradiation of grapes caused a significant increase in anthocyanins content of wine.

248

At the maximal dose of 2700 Gy the highest concentration of total anthocyanins was

249

produced. Interestingly, at the dose of 1300 Gy a minimal concentration of anthocyanins

250

was determined, smaller than in the non-irradiated sample. Similar increases of anthocyanin

251

content in Cabernet and Shiraz wines due to gamma irradiation were reported earlier (Gupta

252

et al., 2015). According to these authors an increase of anthocyanins was proportional to the

253

irradiation dose up to 1500 Gy, and then a decrease was observed at 2000 Gy what was not

254

confirmed in this study. The high dose should enable the beneficial extraction of coloring

255

matter. This more successful extraction was attributed to the increased membrane and cell

256

wall degradation, as well as breakdown of the vacuole membrane of hypodermal cells, thus

257

increasing the release of phenolic compounds in wine (Gupta et al., 2015). Some authors

258

also confirmed the same phenomenon earlier with irradiation of grape pomace (Ayed at al.,

259

1999). Contrary to these results, the study of Alighourchi et al. (2008) showed instability of

260

anthocyanins and their decrease due to gamma irradiation of pomegranate juice especially at

261

doses higher than 2000 Gy. Individual anthocyanins have shown different sensitivity to

262

irradiation. Peonidin-3-O-glucoside was more stable and not strongly influenced by

263

irradiation. However, delphinidin-3-O-glucoside, petunidin-3-O-glucoside and malvidin-3-

264

O-glucoside were more affected. The latter as most representative anthocyanin in V. vinifera

AC C

EP

TE D

M AN U

SC

RI PT

240

12

ACCEPTED MANUSCRIPT grapes increased significantly (p<0.05) with the increase of the radiation dose and reached

266

189.71 mg/L at 2700 Gy, which corresponds to the concentration increase of 20% compared

267

to the concentration in the non-irradiated sample. Similar increase in concentration for

268

malvidin-3-O-glucoside (18 %) in Shiraz wine at lower radiation dose of 1500 Gy was

269

reported by Gupta et al. (2015).

270

Within flavonol compounds an expressive difference was not observed. Some individual

271

flavonols like myricetin-3-glucoside and kaempferol were determinated only in irradiated

272

samples. It seems that gamma irradiation induces releasing soluble phenolic compounds

273

resulting in increased extraction yields (Alothman et al., 2009; Gupta et al., 2015).

274

Quercetin and myricetin were not affected with irradiation treatment. Stability and high

275

resistance of flavonols to gamma irradiation in cranberry syrup was reported earlier,

276

especially for quercetin and myricetin (Rodríguez-Pérez et al., 2015) which are comparable

277

with the results of this study. On the contrary, results of Gupta et al. (2015) showed

278

increased content of quercetin in irradiated samples from Cabernet and Shiraz variety.

279

Hydroxycinnamic and hydroxybenzoic acids showed differences in content due to gamma

280

irradiation but not in same relation to the dose used. Caffeic acid and gallic acid did not

281

significantly change in Merlot wines (p<0.05) regardless to the irradiation dose used. A

282

recent study showed the same results in Shiraz wines where gallic acid and epicatehin

283

remain unchanged due to the irradiation treatment, while an increase of these compounds

284

was observed in Cabernet wines (Gupta et al., 2015). It seems that these differences can be

285

attributed to the grape variety and its botanical characteristics. Concentrations of epicatehin

286

and catehin were not affected with gamma irradiation in Merlot wines.

287

Results of this study have shown that gamma irradiation was a beneficial method only for

288

anthocyanins and the best extractions of phenols were obtained with the highest dose of

AC C

EP

TE D

M AN U

SC

RI PT

265

13

ACCEPTED MANUSCRIPT 2700 Gy, while other phenols like flavonols and flavan-3-ols were not affected by

290

irradiation.

291

3.4 Effect of radiation on volatile aroma compounds of wines

292

Volatile compounds identified in non-irradiated and irradiated wines from Traminer and

293

Merlot variety are shown in Tables 5 and 6. For the better interpretation of the results, mean

294

values of aroma profile data were subjected to principal component analysis. First two

295

principal components explained 84.8 % of the total variance for Traminer wines, and 80 %

296

for Merlot wines. The component patterns and component score plots are shown in Figure 2.

297

The first principal component for Traminer wines is strongly correlated with β-

298

damascenone, nerol, citronellol, cis rose-oxide, β-ionone and geranic acid, and explains 65.5

299

% of the total variance, while α-terpineol contributes more to the second principal

300

component, which explains 19.3 % of the total variance. A component score shows clear

301

segregation of irradiated wines on the upper part of the plot, while the non-irradiated sample

302

is located on the negative side of the first principal component, thus the first component has

303

a stronger discrimination power than the second component. Irradiated wines appear on the

304

upper part of the plane and have greater values of the first component due to higher

305

concentrations of β-damascenone, nerol, citronellol, cis rose-oxide, β-ionone, while non-

306

irradiated wines on the left side of the plane have higher concentration of geranic acid.

307

Thus, irradiation up to 2000 Gy might be beneficial for enhancement of the characteristic

308

aroma of Traminer wine since some of the volatiles like cis rose-oxide have been identified

309

as an important aroma compound in Traminer wines (Robinson, Boss, Solomon, Trengove,

310

Heymann & Ebeler, 2014). The highest dose of 2700 Gy showed higher concentrations of

311

furfural and furfuryl alcohol which can contribute to the toasty and caramel aromas in wine

312

(Pérez-Coello & Díaz-Maroto, 2009).

AC C

EP

TE D

M AN U

SC

RI PT

289

14

ACCEPTED MANUSCRIPT For the Merlot wines the first principal component is strongly correlated with nerol,

314

citronellol, β-ionone and geranic acid, and explains 56.1 % of the total variance, while 2-

315

hexanal, furfuryl alcohol and β-damascenone contribute more to the second principal

316

component, which explains 23.9 % of the total variance. A clear segregation of irradiated

317

samples is evident according to component scores, although the second component showed

318

a more discriminant power. Irradiated wines appear on the right side of the plot in the

319

positive part of the second component, while the non-irradiated sample appears on the

320

negative part of the second component with the higher concentrations of hexanal. Irradiated

321

samples (670 Gy and 1300 Gy) are grouped on the upper part of the plot with the higher

322

concentrations of volatiles like nerol, citronellol, β-ionone and geranic acid, while the

323

sample irradiated with the dose of 2700 Gy is located on the negative part of the first

324

component with the high concentrations of furfural.

325

According to the presented results ionizing irradiation applied in this work showed a

326

positive influence on aroma volatiles in irradiated samples. Overall enhancement in aroma

327

volatiles was observed in wine samples irradiated with doses up to 2000 Gy. An increased

328

content of some volatile compounds due to gamma irradiation application was reported

329

previously (Ananthakumar et al., 2006). Unlike non-irradiated samples, the higher

330

concentrations of benzyl alcohol, phenyl ethyl alcohol and phenyl ethyl acetate were

331

determined in wines from irradiated grapes of Shiraz and Cabernet varieties (Gupta et al.,

332

2015). This phenomenon might be explained by radiation induced breakdown of glycoside

333

precursors (Gupta et al., 2015; Ananthakumar et al., 2006). Increased concentrations of β-

334

damascenone at both varieties due to irradiation can probably be explained by the

335

degradation of carotenes as its precursors (Gupta et al., 2015). Beyond the irradiation dose

336

of 2000 Gy an increase of furfural and furfuryl alcohol concentrations was observed. Results

AC C

EP

TE D

M AN U

SC

RI PT

313

15

ACCEPTED MANUSCRIPT suggest that the highest irradiation dose applied (2700 Gy) enhanced carbohydrate

338

breakdown and caused appearance of furanic derivates in irradiated wines.

339

4. Conclusion

340

The presented study has shown that the use of gamma irradiation suitable for the

341

improvement of phenol and aroma precursor extractions in wine production technology.

342

Irradiation of grapes (var. Traminer and Merlot) had a negative impact on the amino acids

343

content in musts. This should be considered prior to the fermentation process because amino

344

acids are very important nutrients for the yeasts metabolism and allow healthy onset of

345

fermentation. However, wines produced from irradiated grapes showed no lack of quality

346

considering basic chemical composition. Thereby the volatile acidity was generally lower at

347

the higher irradiation doses compared to control wines. According to our results better

348

extraction of the main compounds responsible for the color of red wines demonstrated

349

gamma irradiation to be a promising step in the wine production process. The highest dose

350

of irradiation (2700 Gy) has shown the best results with respect to the concentrations of

351

anthocyanins in Merlot wines. Other phenols like flavonols and flavanols were not affected

352

by irradiation. Aroma profile of the wines prepared from irradiated grapes, especially fruity-

353

floral compounds monoterpens and C13 norisoprenoids, was enhanced with irradiation

354

doses up 2000 Gy. At the highest dose of 2700 Gy more toasty and caramel notes were

355

expressed due to higher concentrations of furfural and furfuryl alcohols. Results in this work

356

indicate that application of gamma radiation could be a feasible and suitable method for the

357

treatment of grapes since in addition of its well-known microbial decontamination

358

efficiency better chemical properties in wine could be achieved.

AC C

EP

TE D

M AN U

SC

RI PT

337

359 360

The authors have declared no conflict of interest.

16

ACCEPTED MANUSCRIPT This research did not receive any specific grant from funding agencies in the public,

362

commercial, or not-for-profit sectors.

AC C

EP

TE D

M AN U

SC

RI PT

361

17

ACCEPTED MANUSCRIPT 363 364

References 1. Abu, J. O., Muller, K., Duodu, K. G., & Minnaar, A. (2005). Functional properties of cowpea (Vigna unguiculata L. Walp) flours and pastes as affected by γ-

366

irradiation. Food Chemistry, 93(1), 103-111.

RI PT

365

2. Alighourchi, H., Barzegar, M., & Abbasi, S. (2008). Effect of gamma irradiation on

368

the stability of anthocyanins and shelf-life of various pomegranate juices. Food

369

Chemistry, 110(4), 1036-1040.

370

SC

367

3. Alothman, M., Bhat, R., & Karim, A. A. (2009). Effects of radiation processing on phytochemicals and antioxidants in plant produce. Trends in Food Science &

372

Technology, 20(5), 201-212.

373

M AN U

371

4. Ananthakumar, A., Variyar, P. S., & Sharma, A. (2006). Estimation of aroma glycosides of nutmeg and their changes during radiation processing. Journal of

375

Chromatography A, 1108(2), 252-257.

376

TE D

374

5. Ayed, N., Yu, H. L., & Lacroix, M. (1999). Improvement of anthocyanin yield and shelf-life extension of grape pomace by gamma irradiation. Food research

378

international, 32(8), 539-543.

380 381

6. Ayed, N., Yu, H. L., & Lacroix, M. (2000). Using gamma irradiation for the

AC C

379

EP

377

recovery of anthocyanins from grape pomace. Radiation physics and chemistry, 57(3), 277-279.

382

7. Bamidele, O.P. & Akanbi, C.T. (2015) Effect og gamma irradiation on amino acids

383

profile, minerals and some vitamins content in pigeon pea (Cajanus Cajan) fluor.

384

British journal of applied science and technology 5(1), 90-98.

18

ACCEPTED MANUSCRIPT 385

8. Caldwell, C. L., & Spayd, S. E. (1989). Effects of gamma irradiation on chemical

386

and sensory evaluation of Cabernet Sauvignon wine. American Chemical Society,

387

USA, 26, 337-345.

389 390

9. Chang, A. C. (2003). The effects of gamma irradiation on rice wine maturation. Food Chemistry, 83(3), 323-327.

RI PT

388

10. Delsart, C., Ghidossi, R., Poupot, C., Cholet, C., Grimi, N., Vorobiev, E., Milisic, V. & Peuchot, M. M. (2012). Enhanced extraction of valuable compounds from merlot

392

grapes by pulsed electric field. American journal of Enology and Viticulture, 63(2),

393

205-211.

M AN U

394

SC

391

11. Fan, X. (2005). Antioxidant capacity of fresh-cut vegetables exposed to ionizing

395

radiation. Journal of the Science of Food and Agriculture, 85, 995-1000.

396

12. Farkas J. (1998). Irradiation as a method for decontaminating food. A review.

400 401 402 403 404 405

TE D

399

13. Garrido, J., & Borges, F. (2013). Wine and grape polyphenols—A chemical perspective. Food research international, 54(2), 1844-1858.

EP

398

International Journal of Food Microbiology, 44, 189-204.

14. González-Neves, G., Favre, G., Gil, G., Ferrer M., & Charamelo, D. (2015) Effect of cold pre-fermentative maceration on the color and composition of young red wines

AC C

397

cv. Tannat. Journal of Food Science and Technology, 56(6), 3449-3457.

15. Gupta, S., Padole, R., Variyar, P. S., & Sharma, A. (2015). Influence of radiation processing of grapes on wine quality. Radiation Physics and Chemistry, 111, 46-56.

16. Harrison, K., & Were, L. M. (2007). Effect of gamma irradiation on total phenolic

406

content yield and antioxidant capacity of Almond skin extracts. Food Chemistry,

407

102, 932-937.

19

ACCEPTED MANUSCRIPT 408

17. Huang, S. J., & Mau, J. L. (2006). Antioxidant properties of methanolic extracts

409

from Agaricus blazei with various doses of g-irradiation. LWT - Food Science and

410

Technology, 39, 707-716. 18. Kunkee, R.E. (1991). Relationship between nitrogen content of must and sluggish

412

fermentation. Proceedings of International Symposium on Nitrogen in Grapes and

413

Wine, Seattle, Davis, 148-155.

414

RI PT

411

19. Lopez, R., Aznar, M., Cacho, J. & Ferreira, V. (2002). Determination of minor and trace volatile compounds in wine by solid-phase extraction and gas chromatography

416

with mass spectrometric detection. Journal of Chromatography A, 966(1), 167-177.

M AN U

417

SC

415

20. Maity, J. P., Chakraborty, S., Kar, S., Panja, S., Jean, J. S., Samal, A. C., Chakraborty, A. & Santra, S. C. (2009). Effects of gamma irradiation on edible seed

419

protein, amino acids and genomic DNA during sterilization. Food Chemistry,

420

114(4), 1237-1244.

TE D

418

21. Moreno-Arribas, M.V. & Polo, P.C. (2009). Amino Acids and Biogenic Amines. In

422

M. V. Moreno-Arribas, & M. C. Polo (Eds.), Wine chemistry and biochemistry (pp.

423

163-191). New York, Springer.

425 426 427 428

22. Nadeem Chandry, M.T. & Evans, R.A. (1971). The effect of γ-irradiation on the

AC C

424

EP

421

amino acid content of proteins. Biochemical Journal, 124 (2), 29-30.

23. O.I.V. (2007). Compendium of International Methods of Wine and Must Analysis. Vol. 1. O.I.V., Paris.

24. Pérez-Coello, M. S., & Díaz-Maroto, M. C. (2009). Volatile compounds and wine

429

aging. In M. V. Moreno-Arribas, & M. C. Polo (Eds.), Wine chemistry and

430

biochemistry (pp. 295-311). New York, Springer.

20

ACCEPTED MANUSCRIPT 431

25. Pripis-Nicolau, L., de Revel, G., Marchand, S., Anocibar Beloqui, A. & Bertrand A.

432

(2001). Automated HPLC method for the measurement of free amino acids including

433

cysteine in musts and wines; first application. Journal of the Science of Food and

434

Agriculture., 81 (8), 731-738. 26. Puértolas, E., Hernández-Orte, P., Sladaña, G., Álvarez, I., & Raso, J. (2010).

RI PT

435

Improvement of winemaking process using pulsed electric fields at pilot-plant scale.

437

Evolution of chromatic parameters and phenolic content of Cabernet Sauvignon red

438

wines. Food Research International, 43(3), 761-766.

SC

436

27. Ražem, D., Anđelić, Lj. & Dvornik, I. (1985). Consistency of ethanol-chlorobenzene

440

dosimetry. Proceedings of IAEA Symposium on High-Dose Dosimetry, Vienna,

441

143-156.

442

M AN U

439

28. Robinson, A. L., Boss, P. K., Solomon, P. S., Trengove, R. D., Heymann, H., & Ebeler, S. E. (2014). Origins of grape and wine aroma. Part 1. Chemical components

444

and viticultural impacts. American Journal of Enology and Viticulture, 65(1), 1-24.

445

TE D

443

29. Rodríguez-Pérez, C., Quirantes-Piné, R., Contreras, M. D. M., Uberos, J., Fernández-Gutiérrez, A., & Segura-Carretero, A. (2015). Assessment of the stability

447

of proanthocyanidins and other phenolic compounds in cranberry syrup after

448

gamma-irradiation treatment and during storage. Food chemistry, 174, 392-399.

450 451 452

AC C

449

EP

446

30. Sacchi, K. L., Bisson, L. F., & Adams, D. O. (2005). A review of the effect of winemaking techniques on phenolic extraction in red wines. American Journal of Enology and Viticulture, 56(3), 197-206. 31. Song, H. P., Kim, D. H., Jo, C., Lee, C. H., Kim, K. S., & Byun, M. W. (2006).

453

Effect of gamma irradiation on the microbiological quality and antioxidant activity

454

of fresh vegetable juice. Food Microbiology, 23, 372-378. 21

ACCEPTED MANUSCRIPT 455 456 457

32. Spinks, J.W.T. & Woods, R.J. (1990). An Introduction to radiation chemistry, 3rd Ed. A Wiley-Interscienece publication, John Wiley&Sons, Inc. New York. 33. Tomaz, I. & Maslov L. (2016). Simultaneous Determination of Phenolic Compounds in Different Matrices using Phenyl-Hexyl Stationary Phase. Food Analytical

459

Methods, 9(2), 401-410.

460

RI PT

458

34. Variyar, P. S., Limaye, A., & Sharma, A. (2004). Radiation-induced enhancement of antioxidant contents of soybean (Glycine max Merrill). Journal of Agricultural and

462

Food Chemistry, 52(11), 3385-3388.

SC

461

35. WHO (1997). High-dose irradiation: wholesomeness of food irradiated with doses

464

above 10 KGy., Joint FAO/IAEA/WHO study group. Technical Report Series, No

465

890, Geneva.

TE D

and Production. New York, Kluwer Academic Publisher.

EP

467

36. Zoecklein, B.W., Fugelsang, K.C., Gump, B.H., Nury, F.S. (1999). Wine Analysis

AC C

466

M AN U

463

22

ACCEPTED MANUSCRIPT Table 1. Chemical composition of non-irradiated and irradiated wines. Data are expressed as the mean ± standard deviation. Control

670 Gy

1300 Gy

2000 Gy

2700 Gy

Ethanol (%vol.)

14.74±0.5 a

14.47±0.33 a

14.74±0.12 a

14.83±0.13 a

15.01±0.45 a

Reducing sugars (g/L)

3.00±0.10 c

2.90±0.06 c

5.80±0.07 a

4.00±0.10 b

4.40±0.05 b

pH

3.01±0.1 a

3.09±0.1 a

3.00±0.1 a

3.04±0.1 a

3.03±0.1 a

Titratable acidity (g/L)*

6.10±0.1 a

5.80±0.1 a

6.20±0.05 a

6.20±0.01 a

6.10±0.02 a

Volatile acidity (g/L)**

0.62±0.02 a

0.60±0.01 a

0.56±0.03 b

0.62±0.02 a

0.52±0.03 b

Sugar free extract (g/L)

18.10±0.05 b

16.90±0.1 c

18.10±0.05 b

Ethanol (%vol.)

13.65±0.48 a

13.39±0.27 a

13.21±0.13 a

13.65±0.5 a

14.02±0.26 a

Reducing sugars (g/L)

1.70±0.06 b

2.10±0.05 a

1.70±0.05 b

2.20±0.07 a

2.10±0.1 a

pH

3.78±0.1 a

3.71±0.1 a

3.65±0.1 a

3.68±0.1 a

3.58±0.1 a

Titratable acidity (g/L)*

4.90±0.05 a

5.00±0.1 a

4.90±0.1 a

4.90±0.05 a

5.10±0.05 a

Volatile acidity (g/L)**

0.60±0.03 b

0.67±0.02 b

0.56±0.04 c

0.74±0.01 a

0.43±0.03 d

Sugar free extract (g/L)

20.70±0.1 a

18.70±0.08 c

19.90±0.05 b

18.10±0.1 c

19.50±0.05 b

SC 18.60±0.06 a

TE D

M AN U

Merlot wine

RI PT

Traminer wine

18.50±0.08 a

AC C

EP

Values with different letters in the same row are significantly different according to the Tukey test (p<0.05). n=3. *expressed as g/L of tartaric acid; **g/L of acetic acid

ACCEPTED MANUSCRIPT Table 2. Amino acids composition of non-irradiated and irradiated Traminer musts. Data are expressed as the mean ± standard deviation.

Control

670 Gy

1300 Gy

2000 Gy

2700 Gy

Aspartate

55.68±1.16 a

20.79±0.53 b

13.19±0.18 c

13.24±0.21 c

10.91±1.57 c

Glutamate

214.12±2.02 a

113.81±3.37 b

89.37±0.78 c

101.97±2.21 bc

90.59±12.31bc

Cysteine

11.87±0.79 a

5.90±0.12 bc

5.20±0.70 c

8.10±0.007 b

8.27±0.98 b

Serine

37.07±0.57 a

19.07±0.45 bc

15.43±0.50 c

21.76±0.36 b

19.73±2.86 bc

Glycine

170.81±0.90 a

73.20±1.85 c

54.27±2.52 d

106.22±0.85 b

79.94±7.13 c

Threonine

45.50±0.26 a

24.87±0.52 c

17.74±0.60 d

30.81±1.06 b

26.41±2.53 bc

Arginine

163.86±2.46 a

89.71±2.65 c

47.13±2.36 d

Alanine

47.27±2.24 a

28.59±0.82 bc

25.13±1.0 c

Tyrosine

10.27±0.1 a

1.14±0.1 b

1.66±0.32 b

Valine

0.48±0.20 a

n.d.

Methionine

24.79±0.31 a

Phenylalanine

SC

RI PT

Amino acids (mg/L)

90.93±11.67 c

40.93±0.44 a

38.85±6.02 ab

4.38±0.42 b

5.46±2.79 ab

n.d.

0.40±0.06 a

0.20±0.28 a

16.97±0.30 c

13.55±0.14 d

22.23±0.12 ab

19.45±1.59 bc

2.09±0.007 ab

0.93±0.14b c

0.11±0.01 c

2.85±0.06 a

1.84±0.73 ab

Isoleucine

6.57±0.13 b

5.85±0.14 b

3.56±0.02 c

9.19±0.22 a

6.27±0.89 b

Leucine

10.16±0.14 a

6.68±0.18 bc

4.15±0.09 c

11.06±0.007 a

8.49±1.62 ab

Lysine

58.36±2.32 a

54.26±3.70 a

48.54±1.11 a

50.68±2.72 a

58.87±10.95 a

TE D

M AN U

139.27±1,37 b

AC C

EP

Values with different letters in the same row are significantly different according to the Tukey test (p<0.05). n=3. n.d. – not detected.

ACCEPTED MANUSCRIPT Table 3. Amino acids composition of non-irradiated and irradiated Merlot musts. Data are expressed as the mean ± standard deviation.

Control

670 Gy

1300 Gy

2000 Gy

2700 Gy

Aspartate

45.18±3.06 a

32.56±0.76 bc

38.40±0.70 ab

24.95±0.11 c

35.23±2.89 b

Glutamate

95.05±7.38 a

78.83±2.92 ab

87.03±1.43 ab

59.22±0.7 c

73.69±7.06 bc

Cysteine

7.02±0.36 a

4.38±0.04 c

6.14±0.13 ab

4.20±0.04 c

5.54±0.41 b

Serine

33.31±2.76 a

28.82±0.71 ab

34.88±0.82 a

23.52±0.05 b

29.76±2.99 ab

Glycine

64.03±3.57 a

54.11±1.36 bc

61.26±0.62 ab

41.08±0.09 d

50.97±3.66 c

Threonine

20.43±1.31 ab

17.99±0.31 bc

22.58±0.40 a

14.86±0.007 c

16.81±1.27 c

Arginine

56.59±3.88 a

41.46±1.18 b

54.27±1.74 a

Alanine

27.19±2.59 ab

22.33±0.66 bc

30.37±0.63 a

Tyrosine

8.31±2.36 a

7.93±0.67 a

10.41±1.09 a

Methionine

21.78±0.81 ab

20.17±0.21 b

Isoleucine

5.38±0.26 b

5.10±0.07 b

Leucine

10.91±0.84 b

Lysine

32.74±3.54 a

SC

RI PT

Amino acids (mg/L)

31.58±3.25 c

19.19±0.03 c

23.76±2.31 bc

6.76±0.14 a

8.18±1.97 a

24.27±0.53 a

16.59±0.17 c

20.62±1.01 b

6.52±0.21 a

4.03±0.04 c

5.37±0.02 b

10.58±0.15 b

12.95±0.42 a

8.54±0.05 c

10.92±0.31 b

37.27±0.09 a

18.08±1.96 bc

16.07±0.48 c

27.33±4.02 ab

a

M AN U

26.61±0.26 c

AC C

EP

TE D

Values with different letters in the same row are significantly different according to the Tukey test (p<0.05). n=3.

ACCEPTED MANUSCRIPT Table 4. Phenolic composition of non-irradiated and irradiated Merlot wines. Data are expressed as the mean ± standard deviation.

670 Gy

1300 Gy

2000 Gy

2700 Gy

Delphinidin-3-Oglucoside

2.86±0.01 d

4.27±0.03 c

2.63±0.005 e

6.03±0.03 b

7.33±0.16 a

Petunidin-3-Oglucoside

10.43±0.05 d

12.96±0.03 c

8.92±0.18 e

14.67±0.2 b

16.81±0.13 a

Peonidin-3-Oglucoside

0.29±0.03 b

0.37±0.03 b

0.28±0.007 b

0.54±0.04 a

0.55±0.06 a

Malvidin-3-Oglucoside

151.74±0.49 d

165.73±0.32 c

137.04±0.74 e

177.76±0.91 b

189.71±0.93 a

∑ Anthocyanins (mg/L)

165.32

183.33

148.87

Myricetin-3glucoside

n.d.

2.41±0.02 b

Myricetin

1.88±0.03 bc

Quercetin

SC

RI PT

Control

214.4

2.00±0.008 c

2.52±0.08 a

2.52±0.01 a

2.12±0.02 a

1.84±0.02 c

2.12±0.03 a

1.97±0.04 b

1.68±0.71 a

3.31±1.68 a

1.34±0.44 a

3.53±1.63 a

3.29±1.49 a

Kaempferol

n.d.

0.44±0.07 a

n.d.

0.29±0.02 b

0.31±0.03 b

Isoramnetin

0.59±0.003 ab

0.79±0.14 a

0.43±0.02 b

0.61±0.16 ab

0.57±0.14 ab

∑ Flavonols (mg/L)

4.15

9.08

5.61

9,07

8.66

Caftaric acid

24.84±0.02 ab

21.81±0.02 c

18.01±0.53 d

25.02±0.49 a

24.05±0.35 b

Caffeic acid

4.18±0.12 a

3.59±0.11 a

3.75±0.39 a

4.25±0.11 a

4.10±0.5 a

Coutaric acid

6.44±0.004 b

6.47±0.01 b

5.71±0.32 c

8.91±0.17 a

9.38±0.44 a

∑Hydroxycinamic acids (mg/L)

35.46

31.87

27.47

38.18

37.53

4.16±0.09 b

4.24±0.07 b

3.70±0.08 b

5.08±0.1 a

3.95±0.42 b

1.20±0.003 c

n.d.

2.73±0.03 b

n.d.

2.91±0.03 a

∑Hydroxybenzoic acids (mg/L)

5.36

4.24

6.43

5.08

6.86

Procyanidin B1

192.36±2.04 a

193.68±2.65 a

191.48±2.26 a

178.54±1.52 b

179.50±3.26 b

Epigallocatehin

13.19±0.01 c

13.51±0.08 bc

13.18±0.07 c

14.97±0.21 a

13.93±0.31 b

Catehin

12.72±0.37 b

13.28±0.48 ab

12.36±0.29 b

14.27±0.24 a

13.55±0.7 ab

Procyanidin B2

11.80±0.27 bc

12.43±0.30 ab

10.92±0.08 c

12.96±0.24 a

13.10±0.58 a

Epicatehin

7.34±0.02 ab

7.55±0.02 a

7.13±0.23 b

7.45±0.06 ab

7.13±0.23 b

∑Flavan-3-ols (mg/L)

237.41

240.45

235.07

228.19

227.21

TE D

AC C

Protocatehinic acid

EP

Gallic acid

M AN U

199

Values with different letters in the same row are significantly different according to the Tukey test (p<0.05). n=3. n.d. – not detected.

ACCEPTED MANUSCRIPT Table 5. Volatile aroma compounds (µg/L) of non-irradiated and irradiated Traminer wines. Data are expressed as the mean ± standard deviation. RT Compounds (µg/L)

LRI

min

Quantifier ion

Control

670 Gy

1300 Gy

2000 Gy

2700 Gy

m/z 33.19

1216

55

n.d.

n.d.

n.d.

1.40±0.02

n.d.

2-hexen-1-ol

45.90

1340

57

n.d.

4.59±0.05

3.06±0.02

1.71±0.17

3.17±0.31

1-hexanol

43.84

1359

56

n.d.

447.8±17.4

406.82±7.17

463.35±38.61

n.d.

452.39

409.88

466.46

3.17

0.43±0.01

0.50±0.003

0.43±0.009

∑ C6 compounds

RI PT

2-hexen-1-al

43.49

1337

139

n.d.

0.46±0.02

Geranic acid

99.54

2353

69

396.39±10.12

72.65±25.33

162.89±127.16

92.57±4.74

218.67±64.3

Linalool

56.70

1551

71

8.40±0.63

15.95±7.03

13.54±5.98

14.92±0.05

9.52±7.74

α-Terpineol

65.65

1684

59

3.67±0.44

3.91±0.22

3.28±0.12

3.96±0.17

2.48±0.13

Citronellol

69.50

1763

69

441.9±8.29

722.3±289.4

624.7±240.9

828.4±9.95

708.2±154.7

Nerol

71.40

1791

69

436±11

715.9±286.9

619.2±238.8

785.8±59.86

702±153.44

1286.36

1531.17

1424.04

1726.15

1641.3

n.d.

0.29±0.42

0.25±0.36

0.66±0.01

0.73±0.01

1.83±0.7

2.22±0.49

3.11±0.09

3.25±0.09

2.12

2.47

3.77

3.98

M AN U

∑ monoterpens 51.84

1451

96

Furfuryl alcohol

64.06

1644

97

n.d.

β-damascenone

72.13

1850

69

0.12±0.03

1.33±0.29

1.09±0.32

1.17±0.12

0.95±0.41

β-ionone

73.92

1879

121

n.d.

1.5±0.03

1.5±0.01

1.34±0.04

1.25±0.12

EP

∑ furans

TE D

Furfural

SC

Cis rose-oxide

0.12

2.83

2.59

2.51

2.20

20.35±0.94

14.98±6.18

11.44±3.62

16.65±0.5

12.44±4.14

∑ C13 norisoprenoids 83.26

2038

85

AC C

γ-nonalactone

n.d. –not detected

LRI- linear retention index consistent with that found in literature

ACCEPTED MANUSCRIPT Table 6. Volatile aroma compounds of non-irradiated and irradiated Merlot wines. Data are expressed as the mean ± standard deviation.

Control

670 Gy

1300 Gy

2000 Gy

2700 Gy

2-hexanal

1.08±1.53

n.d.

n.d.

n.d.

n.d.

2-hexen-1-ol

3.11±4.39

4.19±1.16

2.54±2.22

3.8±0.71

2.71±0.16

1-hexanol

n.d.

n.d.

n.d.

n.d.

∑ C6 compounds

4.19

4.19

2.54

3.8

Cis rose-oxide

n.d.

n.d.

n.d.

n.d.

Geraniol

0.62±0.88

0.78±0.54

0.82±0.54

n.d.

n.d.

Geranic acid

94.25±133.2

201.4±33.3

137.5±103.8

76.29±41.54

n.d.

Linalool

10.18±4.34

9.9±3.4

9.74±0.3

9.41±0.42

7.89±0.27

α-Terpineol

1.18±1.67

2.92±0.05

Citronellol

828.8±217.3

Nerol

RI PT

Compounds (µg/L)

n.d.

2.71

M AN U

SC

n.d.

1.57±0.12

1.16±0.46

912.5±195.3

950±14.42

811.2±132.1

679.2±125.7

821.5±215.5

904.5±193.6

941.7±14.29

804.1±131

673.2±124.6

∑ monoterpens

1756.53

2032

2041.84

1702.57

1361.45

Furfural

0.3±0.43

0.37±0.2

0.28±0.4

0.29±0.41

0.51±0.12

Furfuryl alcohol

0.58±0.82

1.33±0.17

2.22±0.28

2.09±1.25

1.98±0.98

∑ furans

0.88

β-damascenone

TE D

2.08±0.01

2.5

2.38

2.49

501.5±708.1

838.5±163.3

838±9.32

732.9±20.61

618.9±10.4

β-ionone

0.7±0.99

1.42±0.06

1.16±0.04

1.05±0.03

n.d.

∑ C13 norisoprenoids

502.2

839.92

839.16

733.95

618.9

3.56±0.69

4.77±0.95

3.62±0.67

3.1±0.36

γ-nonlactone

4.2±0.67

AC C

n.d. – not detected

EP

1.7

ACCEPTED MANUSCRIPT Figure captions

2

Figure 1. Total amino acids content in non-irradiated and irradiated musts at the irradiation

3

doses of 670, 1300, 2000 and 2700 Gy. (A) Traminer musts, TRC-control; (B) Merlot

4

musts, MEC-control). Means reported here are in mg/L of must, n=3.

5

Figure 2. Principal component analysis (PCA) for volatile aroma compounds of the

6

Traminer (A) and Merlot (B) wines in the plane defined by the first two principal

7

components.

SC

RI PT

1

8

M AN U

9 10 11

15 16 17 18 19 20 21

EP

14

AC C

13

TE D

12

ACCEPTED MANUSCRIPT 22

(A) 900

700 600 500

RI PT

Total amino acids (mg/L)

800

400 300 200

0 TRC

TR670

TR1300

Sample

24

(B)

300

200

100

26 27 28 29 30 31 32

AC C

MEC

25

TR2700

TE D

400

EP

Total amino acids (mg/L)

500

0

TR2000

M AN U

23

SC

100

ME670

ME1300

Sample

ME2000

ME2700

ACCEPTED MANUSCRIPT 33

(A)

M AN U

SC

RI PT

34

35

(B)

37

AC C

EP

TE D

36

ACCEPTED MANUSCRIPT

Highlights

Gamma irradiation had negative impact on amino acids content in grapes

•

Better extraction of coloring matter was obtained from irradiated grapes

•

Flavonols and flavan-3-ols were not affected by irradiation

•

Aroma profile of wines from irradiated grapes was enhanced

•

The irradiation dose up to 2000 Gy increased fruity-floral aroma in wine

AC C

EP

TE D

M AN U

SC

RI PT

•