Sensory profile, biophenolic and volatile compounds of an artisanal ice cream (‘gelato’) functionalised using extra virgin olive oil

Journal Pre-proof Sensory profile, biophenolic and volatile compounds of an artisanal ice cream (‘gelato’) functionalised using extra virgin olive oil Raffaele Sacchi, Nicola Caporaso, Gian Andrea Squadrilli, Antonello Paduano, Maria Luisa Ambrosino, Silvana Cavella, Alessandro Genovese PII:

S1878-450X(19)30019-8

DOI:

https://doi.org/10.1016/j.ijgfs.2019.100173

Reference:

IJGFS 100173

To appear in:

International Journal of Gastronomy and Food Science

Received Date: 16 February 2019 Revised Date:

21 June 2019

Accepted Date: 30 July 2019

Please cite this article as: Sacchi, R., Caporaso, N., Squadrilli, G.A., Paduano, A., Ambrosino, M.L., Cavella, S., Genovese, A., Sensory profile, biophenolic and volatile compounds of an artisanal ice cream (‘gelato’) functionalised using extra virgin olive oil, International Journal of Gastronomy and Food Science (2019), doi: https://doi.org/10.1016/j.ijgfs.2019.100173. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. 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. © 2019 Published by Elsevier B.V.

Sensory profile, biophenolic and volatile compounds of an artisanal ice cream (‘gelato’) functionalised using extra virgin olive oil

Raffaele Sacchi1*, Nicola Caporaso1,2*, Gian Andrea Squadrilli1, Antonello Paduano3, Maria Luisa Ambrosino1, Silvana Cavella1, Alessandro Genovese1

1

Department of Agricultural Sciences, Division of Food Science and Technology, University of Naples Federico II, Portici (NA), Italy

2

Division of Food Science, University of Nottingham, Sutton Bonington, Leicestershire, UK 3

Department of Agricultural and Environmental Science, University Aldo Moro of Bari, Bari, Italy.

*Corresponding authors. E-mail address: [email protected]; [email protected]

1

1

Abstract:

2

This research aimed to characterise an innovative ‘gelato’ (Italian-style artisanal ice

3

cream) made by adding extra virgin olive oil (EVOO) as a functional ingredient, in order to

4

provide health benefits and a characteristic flavour. We report the chemical-physical and

5

sensory profile of an artisanal ‘gelato’ made by adding EVOO (10 % w/w) characterised by

6

medium biophenol content (228 mg kg-1) and a green-herbaceous flavour with a

7

moderately bitter taste. The total phenolic content of the functional EVOO ice-cream was

8

25±0.94 mg kg-1. The additional presence of EVOO added some key volatile compounds,

9

including trans-2-hexenal, 1-hexanol, cis-3-hexen-1-ol and trans-2-hexen-1-ol. The

10

sensory analysis indicated the presence of a slight pungent flavour and “freshly cut grass”

11

aroma, with a slight bitter note given by the EVOO in the ice cream.

12

These findings could lead to the formulation of ice creams in which EVOO is used to

13

partially replace milk fat, with improvement of nutritional profile, and designing new foods

14

with innovative flavours.

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Keywords: Functional foods; gelato; olive oil phenolics; food flavour; SPME-GC/MS; oleic

17

acid.

18

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1. Introduction

20 21

Ice cream is one of the most consumed dairy products worldwide. However, it is

22

generally poor in functional ingredients, and it should be consumed moderately due to its

23

high content in simple sugars and lipids, which are also abundant in short-chain saturated

24

fatty acids (Kurt and Atalar, 2018). Gelato is the Italian-style ice cream, which differs from

25

other ice creams for having little or no overrun, reduced amount of stabilisers and

26

emulsifiers, and generally lower in fat compared to other ice creams (Marshall et al.,

27

2013). In addition, it uses fewer eggs or no eggs; it is also denser as it is churned at a

28

lower speed thus incorporating less air.

29

The interest in functional foods and new eating experiences, e.g. innovative flavours

30

(Van Kleef, Van Trijp, Luning, Jongen, & 2002; Williams, Stewart-Knox, & Rowland, 2005),

31

has been expanding over the years and it has fostered the design of new formulations to

32

improve the nutritional properties of ice cream by using ingredients with enhanced health

33

benefits, e.g. natural antioxidants, colorants, vitamins, proteins, low fat formulations and

34

fats with better fatty acids composition (Prindiville, Marshall, & Heymann, 1999; Prindiville,

35

Marshall, & Heymann, 2000; Welty, Marshall, Grün, & Ellersieck, 2001; Frøst, Heymann,

36

Bredie, Dijksterhuis, & Martens, 2005; Liou & Grün, 2007; Hwang, Shyu, & Hsu, 2009;

37

Choo, Leong, & Henna Lu, 2010; Soukoulis, Lebesi, & Tzia, 2009; Shaviklo, Thorkelsson,

38

Sveinsdottir, & Rafipour, 2011; Sun-Waterhouse, Edmonds, Wadhwa, & Wibisono, 2013;

39

Çam, Erdoğan, Aslan, & Dinç, 2013; ; Karazhian & Mahdian, 2013; Soukoulis, & Tzia,

40

2018; Akalın et al., 2018; Akdeniz & Akalın, 2019).

41

Some small manufacturer of artisanal gelato in Italy have proposed new formulations

42

for this product by including extra virgin olive oil (EVOO), due to its proved sensory and

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nutritional properties (Boskou, 2008; Servili et al., 2009). EVOO can be considered as the

44

top product among vegetable oils, and it is often regarded as a ‘functional ingredient’ due

3

45

to its health benefits given by the presence of phenolic antioxidants, tocopherols,

46

squalene, phytosterols, as well as a ‘natural flavouring agent’ due to its characteristic

47

sensory properties (Boskou, 2008; Sacchi et al., 2014). EVOO adds unique sensory notes

48

due to the presence of hundreds of volatile compounds, but it is also characterised by a

49

typical bitter and pungent taste given by seicoiridoid phenolic compounds (Servili et al.,

50

2009), also referred to as “olive biophenols” (Sivakumar & Uccella, 2010) and

51

characterized by well-known healthy properties (Vitaglione et al., 2015).

52

Olive oil is rich in oleic acid, beneficial for the human health (Boskou, 2015). Health

53

claims have been approved by the European Food Safety Authority for its effect on the

54

maintenance of normal blood cholesterol levels (EFSA, 2011b). In addition, the EFSA

55

approved other health claims related to i) the substitution of saturated fatty acids with

56

mono-unsaturated and/or polyunsaturated fatty acids, which has demonstrated benefits for

57

the human health (EFSA, 2011c) and ii) the health properties of olive biophenols (“Olive oil

58

polyphenols contribute to the protection of blood lipids from oxidative stress”), when the

59

biophenol intake is above a minimum threshold (EFSA, 2011a). Thus, EVOO could be a

60

valuable ingredient for the development of new ice cream products with innovative flavours

61

and enhanced nutritional profile.

62

However, milk proteins are among the major constituents of ice cream, and they are

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expected to interact with EVOO aroma compounds, in an extent depending on the

64

chemical properties of the volatile compounds and the nature of the protein (Guichard,

65

2002; Meynier, Rampon, Dalgalarrondo, & Genot, 2004). EVOO phenolic compounds

66

have also been reported to interact with some food proteins (Pripp, Vreeker, & van

67

Duynhoven, 2005; Genovese, Caporaso, De Luca, Paduano, & Sacchi, 2015), thus

68

lowering its bitterness (Pripp, Busch, & Vreeker, 2004). A strong bitter taste could limit the

69

use of EVOO as an ingredient in many food products because of the low consumers’

70

acceptance (Mc Ewan, 1994; Vitaglione et al., 2013), there could be a potential benefit in

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71

using bitter EVOO in ice cream, with an expected lowering of the EVOO bitter-pungent

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note in this food system with, at the same time, which allows high intake of functional

73

biophenols.

74

This research aimed to verify the sensory and nutritional quality of an Italian artisanal

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ice cream made by using EVOO as its key ingredient. For this purpose, an artisanal ice

76

cream was manufactured by adding an EVOO characterised by a medium content of

77

biophenols, a green-herbaceous flavour and a medium bitter taste. The EVOO-ice cream

78

was then analysed to assess its sensory profile and chemical composition, particularly its

79

volatile and phenolic compounds, by comparing it to a control ice cream made following

80

the same procedure but without EVOO addition.

81 82

2. Materials and methods

83

2.1 Samples, chemicals and reagents

84

Gelato samples were produced by an artisanal ice-cream laboratory (Vanilla Ice Lab,

85

Maddaloni, Caserta, Italy). They were stored at -18 °C and used within a week. A

86

Protected Designation of Origin (PDO) “Colline Salernitane” EVOO was supplied by

87

Torretta olive mill (Battipaglia, Salerno). The oil was obtained from three olive varieties

88

(Carpellese 50%, Frantoio 40% and Rotondella 10%) and its quality was checked

89

according to the European standards (Reg. EU 2568/91 and its subsequent amendments).

90

Additionally, its phenolic composition was analysed by HPLC-DAD, volatile organic

91

compounds by SPME-GC/MS and antioxidant activity by the ABTS method, as detailed

92

below.

93

The reagents and standards used for the analyses were the following: hexane (95%),

94

methanol (99.9%), glacial acetic acid, diethyl ether, ethanol and distilled water were

95

provided by Romil (Cambridge, England). Potassium iodide and sodium carbonate were

96

supplied by AppliChem (Darmsdadt, Germany). Sodium hydroxide, phenolphthalein and

5

97

starch solution were supplied by Titolchimica S.P.A (Rovigo, Italy). Sodium thiosulphate

98

and isobutyl acetate (99.8%) were supplied by Fluka (Buchs, Switzerland). Chloroform

99

was provided by LabScan (Dublin, Ireland). Potassium chloride was supplied by

100

Farmalabor (Pozzillo, Italy). ABTS and Trolox were provided by Sigma-Aldrich

101

(Darmsdadt, Germany).

102 103

2.2 Free acidity, peroxide value and ultraviolet absorbances (K232 and K270) of EVOO

104

EVOO samples were analysed to assess their acidity levels, peroxide value (PV), K232,

105

K270, and ∆K, in accordance to the EU official method (EEC No. 2568/91). Acidity was

106

expressed as oleic acid percentage (%); PV was expressed as meq O2 kg–1 oil. For the

107

analysis of spectrophotometric indices, an ultraviolet-visible UV-1601 spectrophotometer

108

(Shimadzu, Kyoto, Japan) was used. All the analyses were performed in triplicate.

109 110

2.3 Extraction and analysis of phenolic compounds of EVOO

111

EVOO phenolic compounds were extracted and analysed as described by Sacchi et al.

112

(2015). The quantification of individual phenolic compounds was carried out by HPLC-UV.

113

A LC-10ADVP Shimadzu HPLC (Kyoto) equipped with a UV–Vis Diode Array detector

114

(Shimadzu mod. SPD-M10AVP, Kyoto) was used for the analysis. The analyses were

115

performed in triplicate.

116 117

2.4 Ice cream preparation

118

The flow chart of the ice-cream manufacturing process applied is reported in Figure 1.

119

The control for ice cream was prepared in a pasteurizer-emulsifier mixing at 4°C:

120

pasteurised milk (66.7% w/w), a milk cream containing 35% of fat (14.0% w/w) and a mix

121

of powders (19.3% w/w) consisting of skimmed milk powder, sucrose, dextrose, atomised

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glucose and thickeners (guar gum and carob flour). In the pasteurizer, the mixture was

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123

heated up to 85°C and then rapidly cooled to 4°C. An ageing phase of about 10 h was

124

carried out, keeping the mixture in slow stirring. During the aeration-freezing phase, the

125

mixture was continuously stirred for 7-8 min at -8±1 °C. Finally, the ice cream was cooled

126

and stored at -18°C. The EVOO-ice cream process, the pasteurized-homogenized mixture

127

was mixed with an addition of 11.7% EVOO (w/w) and dextrose (3.9%), then it followed

128

the same procedure as the control ice cream.

129 130

2.5 Sensory analysis

131

Sensory analysis of the EVOO used in the preparation of the ice cream was performed

132

according to the official method (Regulation EEC No. 2568/91) to assess its market

133

classification. In order to obtain a better description of the green notes detectable in the

134

fruitiness, two additional descriptors were added to the original profile sheet, i.e. ‘green

135

leaf’ and ‘cut grass’.

136

Sensory analysis of the ice cream was carried out by 9 tasters trained for EVOO

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sensory assessment following the official method (Regulation EEC No. 2568/91). Panel

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test was performed at the Department of Agricultural Sciences, University of Naples

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Federico II (Italy). Several training sessions were carried out to define the sensory

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attributes; consensus was reached among the assessors on the following sensory

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attributes to be measured: ‘milk’ i.e. aromatics reminiscent of cow's milk; ‘cream’ i.e.

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aromatics reminiscent of cream or dairy fat; ‘sweetness’ i.e. fundamental taste sensation of

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which sucrose is typical; ‘olive fruity’ i.e. aromatics reminiscent of EVOO; ‘cut grass’ i.e.

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aromatics reminiscent of the green note of cut grass; ‘pungency’ i.e. fundamental taste

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sensation typical of chili pepper; ‘Bitter’ i.e. fundamental taste sensation given by caffeine

146

or quinine; ‘aromatic persistence’ i.e. the duration or continuation of aroma after

147

swallowing; ‘global aromatic intensity’ i.e. the overall intensity of ice cream aroma; ‘colour’;

148

‘meltdown’ i.e. the time required for the product to melt in the mouth when continuously

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149

pressed by the tongue against the palate; ‘viscosity’ i.e. the measure of flow as the product

150

melts on the tongue when pressed between the tongue and the palate; higher viscosity

151

corresponds to higher scores.; ‘fat feel’ i.e. refers to the intensity of the 'oily' or greasy

152

feeling in the mouth when the product is manipulated between the tongue and the palate;

153

perceived fat content; ‘density’ i.e. the degree of compactness of the sample when

154

pressed between the tongue and palate; ‘iciness’ i.e. the immediate perception of crystal-

155

like particles within the sample. Except for ‘olive fruity’, ‘cut grass’ and ‘pungency’

156

attributes, all these descriptors are usually evaluated in ice cream (Thompson, Chambers,

157

& Chambers, 2009). A 10-point intensity scale with a range of 0 (extremely low) to 10

158

(extremely high) was used.

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2.6 Measure of ice cream viscosity

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The viscosity of the melted control and EVOO ice cream samples was determined by a

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rotational rheometer (HAAKE MARS 60, Thermo Scientific, Waltham, MA) equipped with a

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coaxial cylinders tool (internal diameter 25.08 mm; outer diameter 27.2 mm). The flow

164

curves were carried out at 15°C, in the shear rate range 0.001-10 s-1. Three replications

165

for each sample were performed.

166 167

2.7 Extraction and analysis of phenolic compounds of ice cream

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The extraction of phenolic compounds from ice cream was performed according to

169

Pellegrini et al. (2006), slightly modified. One gram of ice cream was mixed with 5 mL pure

170

methanol and mixed for 15 min at room temperature, followed by a centrifugation at 2500

171

rpm for 10 min (ALC, Milan, Italy). The hydro-alcoholic phase was then taken and filtered

172

for the analysis on a 0.22 nm Mimex-GV filters (Millipore, Cork, Ireland).

173

The quantification of individual phenolic compounds was carried out by HPLC-UV

174

analysis of the hydro-alcoholic extracts (Mateos et al., 2001). A LC-10ADVP Shimadzu

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HPLC (Kyoto) equipped with a UV–Vis Diode Array detector (Shimadzu mod. SPD-

176

M10AVP, Kyoto) was used for the analysis. The analyses were performed in triplicate.

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2.8 Dynamic headspace (DHS) solid phase micro-extraction (SPME) and gas chromatography /mass spectrometry (GC/MS) analysis of ice cream

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DHS-SPME-GC/MS was used for the analysis of volatile compounds in ice-cream

181

samples. The volatile compounds were extracted as described by Welty et al. (2001). Five

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grams of sample were added in a 10 mL vial, and 10 µL isobutyl acetate (408 mg kg-1 in

183

water) was used as the internal standard. The ice cream was collected from the centre of

184

the original container, discarding the top 1 cm of the ice cream. After the ice cream was

185

melted at 25°C for 10 min, a small stirring bar and potassium chloride (1.25 g) was added

186

to the vial, which was tightly capped with a polytetrafluoroethylene (PTFE) septum. The

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content of the vial was stirred vigorously for 1 min. The vial was then placed in a 35±1°C

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stirring water bath for 45 min under moderate constant stirring (550 rpm). After 45 min, the

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prepared fibre was inserted through the septum and fully exposed to the headspace for 15

190

min. The SPME device (Supelco Co., Bellefonte, USA) was equipped with a 50/30 µm

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thickness divinyl-benzene/carboxen/polydimethylsiloxane (DVB/CAR/PDMS) fibre coated

192

with 1 cm length stationary phase. This fibre was chosen for best adsorption of the EVOO

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volatile compounds (Cavalli, Fernandez, Lizzani-Cuvelier, & Loiseau, 2003; Genovese,

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Caporaso, Leone, Paduano, Mena, Perez‐Jimenez, & Sacchi, 2018).

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Volatile compounds were analysed by GC coupled with a mass spectrometer according

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to Sacchi et al. (2015). A GC/MS Shimadzu model QP5050A (Kyoto, Japan) was equipped

197

with a Supelcowax-10 capillary column (60 m, 0.32 mm i.d., 0.5 µm thickness) (Supelco

198

Co., Bellefonte, USA). Thermal desorption of volatile compounds was carried out by

199

putting the SPME fibre in the injector for 10 min. Temperature was set at 40 °C for 4 min,

200

followed by an increase of 3.5 °C min-1 up to 240 °C, held for 3 min. The injector was kept

9

201

at 230 °C. Helium was used as carrier gas (1.4 mL min-1). Volatile compounds thermal

202

desorption was carried out by exposing SPME fibre in the injector for 10 min. The

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compound identification was performed by comparing retention times and mass spectra

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obtained by analysing pure reference compounds in the same conditions. The

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identification was further confirmed by comparing mass spectra with those of NIST

206

database. Mass spectra were recorded at 70 eV. In absence of standard compounds, a

207

tentative identification was reported. The source temperature was 200 °C and the interface

208

temperature was 230 °C. Quantification was given in relation to the internal standard. Peak

209

areas were calculated by using Labsolution acquisition system (GC-MS Solution version

210

1.20; Shimadzu, Kyoto). Before its use, the fibre was conditioned at 270 °C for 1 h for the

211

analysis. A blank test was performed prior to each analysis to prevent the release of

212

undesirable compounds. All the analyses were performed in triplicate.

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2.9 Statistical analysis

215

The statistical significance of the observed differences among ice cream samples was

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assessed for each chemical component by performing a one-way ANOVA. Tukey's test

217

was used to discriminate among the means of the variables. Differences with p < 0.05

218

were considered significant. The data elaboration was carried out using XLStat (version

219

2014.5.03), an add-in software package for Microsoft Excel (Addinsoft Corp., Paris,

220

France).

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3. Results and discussion

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Table 1 reports the free acidity, peroxide value, ultraviolet indices (K232, K270, ∆K),

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phenolic compounds, and sensory attributes of the EVOO used for the ice cream

225

preparation. Free acidity, peroxide value, K232, K270, and ∆K of all EVOO samples were

226

within the law limits of this market category (EC 2568/91). According to the sensory

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227

analysis, EVOO was characterized by medium-intense olive fruitiness (5.7), bitterness

228

(4.5) and pungency (5.0) and a note green leaf (3.4) prevailing on the note of cut grass

229

(1.9).

230

While EVOO pleasant aroma is due to the presence of volatile compounds arising from

231

the LOX pathway, its bitterness and pungency properties are mostly due to phenolic

232

compounds (Caporale et al., 2004; Kalua et al., 2007; Servili et al., 2009).

233

The amount of total phenolic compounds measured in EVOO by HPLC was 228±0.16

234

mg kg-1. Specifically, tyrosol (Ty) content was 9.8±0.2 mg kg-1, hydoxytyrosol (OHTy)

235

6.9±0.1 mg kg-1, dialdehydic form of elenoic acid linked to hydroxytyrosol (OHTy-EDA or

236

‘oleacein’) 71.2±0.2 mg kg-1, dialdehydic form of elenoic acid linked to tyrosol (Ty-EDA or

237

‘oleocanthal’) 60.3±0.4 mg kg-1, aldehydic form of elenoic acid linked to hydroxytyrosol

238

(OHTy-EA or ‘oleuropein aglycone’) 33.8±0.8 mg kg-1, aldehydic form of elenoic acid

239

linked to tyrosol (Ty-EA or ‘ligstroside aglycone’) 10.0±1.0 mg kg-1, coumaric acid 3.4±0.0

240

mg kg-1 and pinoresinol/acetoxy-pinoresinol (PR) 32.3±0.2 mg kg-1.

241 242

3.1 Sensory profile

243

The results of the sensory analyses of the ice cream samples are shown in Figure 2.

244

The EVOO-ice cream had higher ‘colour’ intensity compared to the control ice cream

245

(Figure 2A). This was explained by the typical colour of the EVOO used, which added a

246

yellow note to the ice cream. The perceived ‘viscosity’ of the EVOO-ice cream was not

247

different compared to the control, while the difference in ‘fat feel’ perception was stronger.

248

Milk fat contributes significantly to the creamy flavour of ice-cream (Marshall et al.,

249

2003). In the EVOO ice cream the sensory perception of ‘milk’ and ‘cream’ aromas was

250

lower compared to the control, explained by the masking effect of the EVOO sensory

251

notes (Figure 2B). This latter ingredient caused the appearance of intense ‘pungent’, ‘olive

252

fruity’ and ‘cut grass’ notes. The ‘green leaf’ note detected in EVOO (Table 1), however,

11

253

was not found in the EVOO ice cream and‘bitterness’ was perceived with a very low

254

intensity (0.8) compared to that of the added EVOO (4.5) (Table 1). The low bitterness

255

perception in the EVOO-ice cream may be related to the presence of sweet compounds,

256

but also to the interaction between EVOO phenolic compounds and milk proteins (Pripp et

257

al., 2004). Phenolic compounds, in fact, are known to interact with food proteins by

258

covalent and non-covalent binding, leading to a decrease of bitterness perception (Pripp et

259

al., 2005; Genovese et al., 2015). EVOO ice cream had a slightly higher “overall aroma”

260

intensity and “persistence” than the control.

261 262

3.2 Measure of viscosity

263

EVOO ice cream had a higher viscosity than the control one. In particular, at shear rate

264

of 1 s-1 EVOO ice cream viscosity was twice (0.044 Pa s) that of the control (0.022 Pa s).

265

This result confirms the perceived sensory difference in terms of viscosity given by the

266

sensory panel.

267 268

3.3 Biophenols

269

The total content of phenolic compounds measured in the EVOO-ice cream was

270

25±0.9 mg kg-1. Specifically, it had 3.9±0.0 mg kg-1 OHTy, 5.8±0.2 mg kg-1 Ty, 4.6±0.0 mg

271

kg-1, 5.5±0.5 mg kg-1 OHTy-EA and 5.1±0.3 mg kg-1 Ty-EA. Compounds such as coumaric

272

acid, Ty-EDA and PR were not detected in the ice cream, despite being found in the initial

273

EVOO. In addition, the ratio between the compounds OHTy-EA and Ty-EDA in the

274

functionalised ice cream was different from the bulk EVOO sample. The content of OHTy-

275

EA was 5.5±0.5 mg kg-1, while PR was not found in the functionalised ice cream, despite

276

the content of these two molecules was similar in the EVOO (32 and 33 mg kg-1). This

277

finding could be related to the selective interaction of these compounds with milk proteins

278

(Pripp et al., 2005) and also explains the different ratio between ‘pungent’ and ‘bitter’

12

279

sensory attributes perceived in EVOO ice cream. The difference in oil/water partition for

280

each biophenolic molecule can also contribute to explain these differences (Fogliano et al.,

281

1999).

282

As far as the nutritional functionality of EVOO ice-cream, considering a portion size for

283

ice cream from 66 to 200 grams, the potential phenolic intake from one portion would be

284

approximately 2-5 mg. This amount ensures to cover from 30 to 100% of the minimum

285

daily intake suggested by the European Food Safety Authority (EFSA) for virgin olive oil to

286

exert its physiological function in protecting blood lipids from oxidative stress (EFSA,

287

2011a). This confirms the potential nutritional usefulness of an ice-cream containing

288

EVOO with a medium level of seicoiridoid biophenols (200-300 mgkg-1) as ingredients.

289

The use of a more ‘intense fruity’ EVOO characterised also by an higher level of

290

biophenols (400-800 mg kg-1), such as those produced in Italy and Greece as monitored

291

by specific projects (ARISTOIL, 2019; Ager-2 COMPETiTiVE, 2017-2019), could allow

292

reaching the minimum intake suggested by the EFSA health claim when consuming a

293

smaller ice-cream portion.

294

The masking effect of EVOO bitterness and pungency likely due to milk protein-

295

biophenol interactions, suggests that increase in EVOO biophenolic intake would be

296

possible in the ice-cream (or other traditional Mediterranean food preparations and

297

recipes) by using the concept of food pairing to enhance acceptability and sensory

298

preference, which is known to be negatively influenced by bitterness (Cavallo et al., 2019).

299 300

3.4. Volatile compounds

301

Volatile compounds quantified in the control and EVOO-ice cream are reported in Table 2,

302

with the indication of their sensory attributes. The most abundant compounds in the control

303

ice cream were 2-butanone, limonene and hexanal. These compounds mostly originate

304

from both the animal metabolism and the type of feed (Cadwallader, & Singh, 2009).

13

305

Hexanal arising both from unsaturated fatty acid degradation in milk (Asaduzzaman,

306

Biasioli, Cosio, & Scampicchio, 2017) and LOX activity in VOO and can be associated to

307

positive sensory ‘green’ notes (Kalua et al., 2007). Volatile compounds in EVOO-ice cream

308

were mostly represented by ‘green odours’ like trans-2-hexenal (which was by far the most

309

abundant compound), 1-hexanol, cis-3-hexen-1-ol and trans-2-hexen-1-ol, whose

310

contribution to the overall volatile profile was intense, as their concentration was above 2.5

311

and 1.5 mg kg-1, respectively, while the other compounds were found at concentrations

312

below 0.5 mg kg-1. The volatile compounds arising from the added EVOO, represent the

313

major contributors to the “green-grass” and “fruity” aroma (Kalua et al., 2007). A

314

contribution to the astringent-bitter taste can be also attributed to trans-2-hexen-1-ol

315

(McEwan, 1994) and cis-3-hexen-1-ol (Caporale et al., 2004). Other compounds

316

previously reported in EVOO include 1-penten-3-one, 1-penten-3-ol, cis-2-hexenal and cis-

317

2-penten-1-ol, which were found in the EVOO-ice cream at relatively high abundance.

318 319

CONCLUSIONS

320

The results of this study demonstrated that the use of EVOO in the formulation of an

321

Italian artisanal ice-cream (‘gelato’) caused significant changes in the volatile profile,

322

phenolic compounds, viscosity and sensory attributes. This innovative EVOO-ice cream

323

could be regarded as a ‘functional food’, due to the improved health benefits arising from

324

the added EVOO, which can be further modulated by the use of EVOOs with higher

325

contents of biophenols.

326

The results here presented also suggest an interaction between the ice-cream matrix

327

and the EVOO components, which could impact the extractability of phenolic compounds

328

but also influence the sensory perception of EVOO bitterness. As bitterness is usually

329

associated to low consumer’s acceptability, this “masking effect” in the ice-cream matrix

330

would be beneficial in order to manufacture products with higher concentration of phenolic

14

331

compounds without the risks of obtaining low consumer acceptability due to EVOO

332

bitterness.

333

The industry could also look to other strategies of adding olive-derivative phenolic

334

compounds such as recovery from the olive mill wastewater to be used in ice creams.

335

Further investigations could also focus on the partial replacement of milk fat with EVOO by

336

keeping constant the amount of total fat, as well as to better understand the mechanisms

337

involved in the interaction between milk proteins and EVOO biophenols in ice cream and

338

their bioavailability.

339 340 341

ACKNOWLEDGMENTS

342

Pina Molitierno (Vanilla Ice lab, Caserta, Italy) and Maria Provenza (Oleificio Torretta,

343

Battipaglia, Italy) are acknowledged for their help in ice-cream and olive oil production,

344

respectively. This research has been supported by Progetto AGER-2 (grant n. 2016-0174)

345

COMPETiTiVE (Claims of Olive oil to iMProvE The market ValuE of the product;

346

www.olivoeolio.progettoager.it).

347

All authors declare no conflict of interest.

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FIGURE CAPTION

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Figure 1 - Flow chart for the manufacturing of the artisanal extra virgin olive oil ice cream (gelato) used in this experiment.

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Figure 2 - Sensory profiles of visual and physical sensations (A), and olfactory-taste

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characteristics (B) of extra virgin olive oil (EVOO) ice cream compared to a blank

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sample without EVOO. Asterisks indicate statistically significant differences (p < 0.05).

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Table 1. Quality indices, phenolic compounds and sensory attributes for the extra virgin olive oil used in ice cream production.

Quality indices Acidity Peroxide value K232 K270 ∆K Phenolic compounds OHTy Ty Cumaric acid OHTy-EDA Ty-EDA PR OHTy-EA Ty-EA Total phenolic compounds Sensory attributes Pungency Bitterness Olive fruity Cut grass Green leaf

EVOO

EVOO law limits*

0.34±0.01 10.70±0.04 2.317±0.110 0.149±0.009 -0.003±0.001

≤ 0.8 < 20 ≤ 2.50 ≤ 0.22 ≤ 0.01

6.9±0.1 9.8±0.2 3.4±0.1 71.2±0.2 60.3±0.4 32.3±0.2 33.8±0.8 10.0±1.0 228.0±0.2

-

5.0 4.5 5.7 1.9 3.4

>0 -

515 516 517 518 519 520 521 522

* EC Reg. 2568/91 and further modifications. Acidity is expressed as oleic acid equivalent (g/100g). Peroxide value is expressed as meq O2 kg-1 oil. Phenolic compounds content is expressed as mg kg-1. Values are the average of three replicates of analysis (n=3). OHTy, hydoxytyrosol; Ty, tyrosol; OHTy-EDA, dialdehydic form of elenoic acid linked to hydroxytyrosol; Ty-EDA, dialdehydic form of elenoic acid linked to tyrosol; PR, pinoresinol/acetoxypinoresol; OHTy-EA, aldehydic form of elenoic acid linked to hydroxytyrosol; Ty-EA, aldehydic form of elenoic acid linked to tyrosol. Sensory attributes are expressed as median on an unstructured 0-10 scale.

523

23

524 525

Table 2. Content of volatile compounds (µg kg-1) in control and EVOO-functionalised ice cream, with indication of their sensory descriptors. Compound

Sensory descriptora

Originb

EVOO ice cream

Sulphurous, M 1.74±0.34 a 2.36±0.30 a vegetable Octane Sweet, alkane M/EVOO 1.49±0.16 a 8.97±0.26 b Ethyl acetate Pungent M/EVOO 6.47±0.59 a 90.72±6.56 b 2-Butanone Ethereal, fruity M 25.91±3.78 a 32.39±2.71 a 3-Methylbutanal Almond M/EVOO 2.54±0.35 a 2.83±0.23 a Ethanol Fruity, sweet M/EVOO 1.65±0.17 a 22.54±1.96 b Propyl acetate Celery M 5.91±0.55 a 5.73±0.44 a Alcohol, apple, 2-Pentanone M 2.31±0.24 b NF a banana, cheese 3-Pentanone Fruity, green, sweet M/EVOO 2.38±0.31 a 136.75±13.00 b 1-Penten-3-one Pungent EVOO NF a 376.65±20.38 b Hexanal Green M/EVOO 13.97±0.62 a 211.70±4.68 b trans-2-Pentenal Grass, bitter, almond EVOO NF a 53.17±4.68 b 1-Penten-3-ol Olive oil, plastic EVOO NF a 439.68±35.67 b 2-Heptanone Sweet, fruity M 7.03±0.96 a 6.65±0.60 a Heptanal Oily, fatty, wood M/EVOO 1.68±0.20 a 2.87±0.23 b 3-Methyl-1-butanol Pungent M/EVOO 0.91±0.10 a 12.68±1.14 b d-Limonene Citrus, floral M/EVOO 14.54±1.88 b 10.48±0.95 a cis-2-Hexenal Green, fruity, sweet EVOO NF a 278.16±10.22 b tran-2-Hexenal Grass EVOO NF a 15645.25±708.8 b 1-Pentanol Pungent EVOO NF a 66.82±1.72 b Hexyl acetate Banana, fruity EVOO NF a 36.90±0.18b Fatty, sharp, citrusOctanal M/EVOO 0.70±0.09 a 10.80±0.67 b like, soapy cis-2-Penten-1-ol Fatty, almond EVOO NF a 370.94±21.47 b cis-3-Hexenyl acetate Herbaceous, Banana EVOO NF a 85.93±4.30 b Apple, banana, trans-2-Hexenyl grape fruity, EVOO NF a 13.15±0.53 b acetate herbaceous trans-2-Heptenal Tallow, pungent EVOO NF a 36.75±3.40 b 1-Hexanol Green, fruity, floral EVOO NF a 2844.71±76.05 b cis-3-Hexen-1-ol Fruity, herbaceous EVOO NF a 1826.97±64.71 b 2-Nonanone Fruity, apple M 1.12±0.09 b 0.36±0.02 a trans-2-hexen-1-ol Fruity EVOO NF a 3922.44±108.07 b Values are the average of three replicates (n=3), followed by the standard deviation. -1 *: M=Milk; EVOO= Extra virgin olive oil; M/EVOO= Milk and EVOO. The content is expressed as µg kg ; for -1 molecules deriving from the milk, it is expressed as µg kg of milk constituents in the ice cream sample, for -1 those deriving from the oil they are µg kg of oil, while for those originated from both from milk and oil they -1 are expressed as µg kg of milk constituents and oil. NF = not found. Different letters indicate significant differences (p < 0.05) Dimethyl sulphide

526 527 528 529 530 531 532

Control ice cream

533

24

Highlights: Italian-style ice cream (gelato) was formulated using extra virgin olive oil The effect of sensory profile, phenolics and volatile compounds was investigated EVOO-gelato had

25±0.9 mg kg-1 phenolics, and volatiles with grass-fruity notes

A possible interaction between EVOO and milk proteins in gelato was suggested

Conflict of interest statements

On behalf of all co-authors, I certify that none of the co-authors of the manuscript submitted have any conflict of interest related to this research.

22.09.2018 Prof. Raffaele Sacchi