The combination of modified atmosphere packaging with eugenol or thymol to maintain quality, safety and functional properties of table grapes

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Postharvest Biology and Technology 41 (2006) 317–327

The combination of modified atmosphere packaging witheugenol or thymol to maintain quality, safety and

functional properties of table grapes

D. Valero a,∗, J.M. Valverde a, D. Martınez-Romero a, F. Guillen a, S. Castillo a, M. Serrano b

a Department of Food Technology, University Miguel Hernandez, Ctra. Beniel km. 3.2, 03312 Orihuela, Alicante, Spainb Department of Applied Biology, University Miguel Hernandez, Ctra. Beniel km. 3.2, 03312 Orihuela, Alicante, Spain

Received 6 October 2005; accepted 13 April 2006

bstract

Active packaging was developed by adding eugenol or thymol to table grapes stored 56 days under modified atmosphere (MAP). Control

erries showed losses of quality in terms of sensory, nutritional and functional properties. These losses were significantly reduced in packagesith added eugenol or thymol. In addition, lower microbial spoilage counts were achieved with the active packaging. Thus, this simple

echnology could be an interesting tool to preserve the overall quality of table grapes.2006 Elsevier B.V. All rights reserved.

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eywords: Active packaging; Antioxidant activity; Essential oils; MAP

. Introduction

The term quality implies a grade of excellence of aroduct and its preparation for a specific use, and involvesrganoleptic, nutritive and functional properties (Abbott,999). Polyphenols are the most important constituents ofrapes from the point of view of functional properties, withatechins, flavonol glycosides, phenolic acids, stilbenes andnthocyanins all having been detected (Threlfall et al., 2005).ll these compounds have been shown to exert antioxidant

ctivity in vitro (Bonilla et al., 1999), and health bene-t effects preventing artheriosclerosis, stroke and coronaryeart disease and cancer, among other diseases (Tomas-arberan and Espın, 2001; Yilmaz and Toledo, 2004). How-ver, changes in anthocyanin profiles and loss of other phe-olic compounds have been reported during advanced stagesf ripening (Canals et al., 2005). In this sense, the ripening

rocess in table grapes leads to a reduction in functional prop-rties, and consumers therefore may not achieve the benefitsrom eating these fruit.

∗ Corresponding author. Tel.: +34 96 6749743; fax: +34 96 6749677.E-mail address: [email protected] (D. Valero).

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925-5214/$ – see front matter © 2006 Elsevier B.V. All rights reserved.oi:10.1016/j.postharvbio.2006.04.011

Table grape is a non-climacteric fruit with severe prob-ems during postharvest handling, storage and marketing.he quality losses are based mainly on weight loss, colourhanges, and accelerated softening. Additionally, deteriora-ion in table grapes is also attributed to rachis browning andigh incidence of berry decay, mainly due to Botrytis cinerea.he classic control of this rot has been the use of the synthetic

ungicide sulphur dioxide (SO2), but the necessary concen-ration may induce injuries in both rachis and berry (Crisostot al., 2002). In addition, sulphite residue is an important con-umer problem, as well as a warning to the consumer abouthemical usage.

To solve this problem, several strategies have been devel-ped, such as storage with high CO2 (Retamales et al.,003), hypobaric treatments (Romanazzi et al., 2001), andiocontrol (Schena et al., 2002), but occurrence of injuriesrachis browning and off-flavours) has occurred with thesereatments. Modified atmosphere packaging (MAP) also pre-erves table grape organoleptic quality (Martınez-Romero

t al., 2003), but the CO2 concentration inside packages issually not high enough to exert a fungicide effect, and aO2-commercial generator is necessary (Artes-Hernandez etl., 2004), and the problem of SO2 still exits.

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18 D. Valero et al. / Postharvest Biol

All these concerns have stimulated the search for newtrategies, such as the use of natural antimicrobial com-ounds, as alternative means for controlling postharvestecay while maintaining table grape quality. For centuries,ssential oils have been empirically recognized as antimicro-ial, and scientific confirmation has been reviewed recentlyBurt, 2004). In this sense, eugenol (clove essential oil com-ound, Syzygium sp.) and thymol (from thyme and oregano,hymus and Origanum) exhibit antioxidant and antimicro-ial properties (Lee and Shibamoto, 2001; Kulisic et al.,004). The antioxidant effect of these compounds has beeneported to be higher than �-tocopherol or BHT (Lee et al.,005; Sacchetti et al., 2005), while their antimicrobial activ-ty has been proven with bacteria (Periago et al., 2004), fungiVazquez et al., 2001) and yeasts (Arora and Kaur, 1999).

In foods, the use of essential oils or some of their individ-al components, as potential natural preservatives has beeneported in cheese (Smith-Palmer et al., 2001), bakery prod-cts (Guynot et al., 2003) and meat (Quintavalla and Vicini,002), although there is some evidence for the control ofruit spoilage. In recent reports, eugenol, menthol or thymoln combination with MAP maintained quality and safety byeducing microbial spoilage in sweet cherry (Serrano et al.,005a) and ‘Crimson’ table grapes (Valverde et al., 2005).he amount added of the pure compounds inside the pack-ges was 1 and 0.5 ml for sweet cherry and table grape,espectively. A slight odour of these compounds was detectedmmediately after opening the packages, which disappearedery rapidly with time due to evaporation at room temper-ture. However, after tasting the grapes, persistence of theharacteristic aroma was detected, especially for menthol,hile eugenol showed the lowest residual flavour.The purpose of this work was to use eugenol or thymol at

uch lower concentrations (75 and 150 �l inside the pack-ges) and to study the effect of these compounds on tablerape quality, safety, and nutritional and functional proper-ies. As far as we know, this is the first paper in which aroad study of the use of eugenol and thymol in combinationf MAP has being carried out in fruit.

. Material and methods

.1. Experimental procedure

Table grapes (Vitis vinifera L. cv. Autumn Royal) werearvested from a commercial farm in Abaran (Murcia,pain). At the laboratory, clusters (150–170 g) were selected

o obtain homogeneous batches based on colour, size,bsence of injuries and healthy, greenish rachises. Clus-ers were packed in 20 �m thickness non-perforated ori-nted polypropylene (N-OPP) bags (30 cm × 20 cm), which

ad permeabilities at 1 ◦C of 1600 ml O2 m−2 d−1 atm−1 and600 ml CO2 m−2 d−1 atm−1. This film was selected basedn previous experiments with table grapes (Martınez-Romerot al., 2003). Treatments with eugenol or thymol (99.5%

2

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Technology 41 (2006) 317–327

urity and purchased from Sigma, Sigma–Aldrich, Madrid,pain), were performed by placing either 75 or 150 �l of theseatural compounds on sterile gauze inside the bag, avoidinghe contact with berries, and then immediately sealing to min-mize vaporization. Clusters packed in the same conditionsut without treatments served as controls. All packages weretored at 1 ◦C and 90% RH in darkness for 56 days. Ten bagsor each treatment were taken after 14, 28, 42 and 56 days,alf of them were immediately analysed (cold storage) andhe remainder transferred to a chamber under controlled con-itions at 20 ◦C and 90% RH and analysed after 2 days, toimulate market operations (shelf life, SL). The followingnalytical determinations were performed at harvest (day 0)nd at each sampling date.

.2. Gas composition

A silicone septum was provided on the bag surfaceor sampling gas inside the package. One millilitre of theeadspace atmosphere was withdrawn using a gas syringe andsed to quantify ethylene, CO2 and O2 concentrations insidehe packages as previously reported (Martınez-Romero et al.,003). Results were expressed as �l l−1 of ethylene and kPa2 and CO2 and were the mean ± S.E. of two determinations

or each of the five replicates (n = 10).

.3. Weight loss, colour and firmness

Weight of individual bunches was recorded on the dayf harvesting and after the different sampling dates. Cumu-ative weight losses were the mean ± S.E. of five bunchesnd expressed as percentage loss of original weight. Colouras determined using the Hunter Lab System and a Minolta

olorimeter CR200TM model (Minolta Camera Co., Osaka,apan). The individual L*, a* and b* parameters wereecorded and results were the mean ± S.E. of determina-ions made on 10 berries for each bag along the equato-ial axis (n = 50). Flesh firmness was determined using aA-XT2i Texture Analyzer (Stable Microsystems, Godalm-ng, UK) interfaced to a personal computer, as previouslyeported (Martınez-Romero et al., 2003). Results were theean ± S.E. of determinations made in 10 berries for each

ag (n = 50) and expressed in N.

.4. Maturity index determination

Total soluble solids concentration (TSS), titratable acid-ty (TA) and maturity index (TSS/TA) were determined inriplicate in the juice obtained from 10 berries for each bags previously reported (Martınez-Romero et al., 2003). Allesults were expressed as the mean ± S.E. (n = 15).

.5. Sensory analysis

Sensory analyses to compare the quality of treated andontrol table grapes were carried out by 10 trained adults,

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D. Valero et al. / Postharvest Biol

ged 25–40 years (five female and five male). The panel wasrained in a pre-test in which berries with extremely low origh attributes (crunchiness, juiciness, sweetness, sournessnd quality) were evaluated (Martınez-Romero et al., 2003).or the rachis, symptoms of dehydration and browning forrimary and secondary branches were evaluated on a rankedcale of 1–5, where 1, absence of these symptoms; 2, slightccurrence; 3, moderate; 4, severe; and 5, extremely severerowning and dehydration. For each treatment and samplingate, the five branches were evaluated by each judge andesults were the mean ± S.E. (n = 50). In a second test, eachudge evaluated five berries for each treatment for the follow-ng characteristics: visual aspect (general aspect), firmness,weetness, juiciness, sourness and crunchiness, on a scalef 1–5 (ranked), where 1, very low; 2, low; 3, medium; 4,igh; and 5, very high. Results were the mean ± S.E. (n = 50).inally, the occurrence of off-flavours (presence or absence)as also tested using a triangle test according to O’Mahony

1986). The sensory analyses were made after 56 days of coldtorage plus 4 days at 20 ◦C.

.6. Decay and microbiological analysis

For decay analysis, the number of decayed and fadedrapes for each bunch was counted, and then percentage ofhe total berries was calculated for each sampling date. After8 and 56 days at 1 ◦C, samples of 10 g from each bag werebtained under sterilized conditions to count mesophilic aer-bics and mould and yeast, as previously reported (Serranot al., 2005a).

.7. Antioxidant activity, total phenolic compounds andotal anthocyanins

Twenty berries from each bag were peeled to obtain skinnd pulp separately. Total antioxidant activity (TAA) and totalhenolics were quantified in skin or pulp phosphate bufferxtracts. Anthocyanins were determined using 2 g of skin tis-ue homogenized in 4 ml methanol by reading the absorbancet 530 nm. All these methods have been detailed previouslynd parameters quantified in duplicate (Serrano et al., 2005b).esults were the means ± S.E. (n = 10).

.8. Organic acids and sugar contents

Organic acid and sugar determinations were quantified inhe same extract as above, by HPLC using absorbance andefractor index detectors, as previously reported (Serrano etl., 2005b). For both, organic acids and sugars, the resultsere the means ± S.E. of determinations made in duplicate

or each bunch (n = 10).

.9. Statistical analysis

Data for the organoleptic, nutritive, microbial, functionalnd sensory parameters were subjected to analysis of vari-

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Technology 41 (2006) 317–327 319

nce (ANOVA). Sources of variation were time of storagend treatments. Mean comparisons were performed usingSD the Tukey’s test to examine if differences between

reatments and storage time were significant at P < 0.05. Allnalyses were performed with SPSS software package v. 11.0or Windows. Linear regressions were performed among theunctional parameters taking into account all sampling dataither during cold storage or shelf life.

. Results

.1. Gas composition inside MAP packages

The CO2 concentration increased and the O2 decreased forll treatments during cold storage. A no-significant slight dif-erence in the final gas composition was observed betweenontrol and treated packages. Thus, control clusters led tohe highest CO2 (≈4 kPa) and the lowest O2 (≈7 kPa) lev-ls inside the packages, while the contrary was detectedor the thymol treatment (≈2.5 and 8–10 kPa for CO2 and

2, respectively). The atmosphere composition reached withugenol-treated clusters ranged in between with concentra-ions of ≈3.3 and ≈6.5 kPaO2. In addition, no significant dif-erences were obtained between 75 and 150 �l doses. Whenlusters were transferred to 20 ◦C (SL), a greater increase inO2 and decrease in O2 was observed with final levels of–7 kPa CO2 and O2 without significant differences amongreatments.

.2. Parameters related to berry quality

The cumulative weight loss increased during storagelthough it was significantly higher in control than inreated bunches. Thus, after 56 days at 1 ◦C, control clus-ers had lost 1.60 ± 0.20%, the losses being greater afterL (2.10 ± 0.18%). At these periods, treated clusters lost.35–0.50% (cold) and 0.8–0.9% (SL), without significantifferences due to treatment or applied dose.

‘Autumn Royal’ table grape is characterised by a purple-lack skin colour at the optimum stage of harvest, at which theolour coordinates were 39.09 ± 0.53 (L*), 3.43 ± 0.36 (a*)nd −2.02 ± 0.19 (b*). During cold storage and SL, controlerries showed the greater variations of colour with signifi-ant decreases in L* and a* parameters, while slight increasesere observed for b* (Table 1). Changes in L* and a* were

educed by the addition of eugenol and especially thymol, theelay being higher with the maximum applied dose (150 �l).

With respect to flesh firmness, an accelerated soften-ng process in control clusters as the storage advanced wasbserved. Thus, values at harvest (4.03 ± 0.15 N) decreasedo levels of 2.49 ± 0.22 and 2.47 ± 0.19 N after 56 days ofold storage and SL, respectively. In contrast, the addition of

ugenol or thymol inside the packages significantly delayedhe loss of firmness with no significant differences dependingn the added compound or applied dose, with final firmnessevels of ≈3.5–3.8 N.

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Table 1Colour parameter (L*, a* and b*) values at harvest and after 56 days of coldstorage or SL (4 days at 20 ◦C) of table grapesa

Parameter Harvest 56 Cold 56 + SL

L*

Control 39.09 ± 0.53 a 25.96 ± 0.61 b A 25.06 ± 0.32 b AE-75 27.93 ± 0.48 b B 27.36 ± 0.54 b BE-150 28.18 ± 0.60 b B 27.95 ± 0.68 b BT-75 26.76 ± 0.49 b A 26.15 ± 0.56 b BT-150 28.30 ± 0.67 b B 27.90 ± 0.61 b B

a*

Control 3.43 ± 0.36 a 1.22 ± 0.17 b A 1.11 ± 0.15 b AE-75 1.43 ± 0.21 b A 1.33 ± 0.14 b AE-150 1.60 ± 0.23 b A 1.41 ± 0.14 b AT-75 2.12 ± 0.41 b B 1.71 ± 0.27 b BT-150 2.27 ± 0.31 b B 1.73 ± 0.35 b B

b*

Control −2.02 ± 0.19 a −1.19 ± 0.11 b A −1.08 ± 0.18 b AE-75 −1.29 ± 0.25 b A −1.33 ± 0.17 b AE-150 −1.41 ± 0.18 b A −1.49 ± 0.14 b AT-75 −1.43 ± 0.29 b A −1.43 ± 0.22 b AT-150 −1.52 ± 0.10 b A −1.52 ± 0.29 b A

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Table 2Levels of TSS (g 100−1) and TA (g equiv. tartaric acid 100 g−1) in tablegrapes at harvest and after 56 days of cold storage or SL (4 days at 20 ◦C)a

Parameter Harvest 56 Cold 56 + SL

TSSControl 18.34 ± 0.16 a 21.10 ± 0.23 b A 21.42 ± 0.21 b AE-75 19.14 ± 0.18 a B 19.59 ± 0.20 b BE-150 19.03 ± 0.13 a B 19.29 ± 0.19 b BT-75 19.32 ± 0.39 a B 19.97 ± 0.19 b BT-150 19.09 ± 0.29 a B 19.54 ± 0.19 b B

TAControl 0.48 ± 0.01 a 0.28 ± 0.01 b A 0.26 ± 0.01 b AE-75 0.31 ± 0.02 b B 0.30 ± 0.02 b BE-150 0.34 ± 0.01 b B 0.32 ± 0.01 b BT-75 0.41 ± 0.02 a C 0.39 ± 0.01 a CT-150 0.43 ± 0.02 a C 0.41 ± 0.01 a C

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or each parameter, similar small letters in rows are not significantly differentt P < 0.05 from levels at harvest and during storage. For each column, similarapital letters are not significantly different at P < 0.05 among treatments.

Levels of TSS and TA at harvest were 18.34 ± 0.16◦ Brixnd 0.48 ± 0.01 g 100 g−1 tartaric acid equivalent, respec-ively, resulting in a maturity index (TSS/TA) of 38.48 ± 1.66.SS/TA showed increases throughout storage, which wereignificantly higher in control than in treated berries. Athe end of cold storage, control fruit showed levels ofSS/TA of 76.05 ± 2.33 which significantly increased afterL (82.97 ± 2.11). On the contrary, treated table grapeselayed the change in maturity index, but differently depend-ng on the added compound. Thus, thymol was more effectivehan eugenol in this delay, although the 150 �l dose gavelightly better results in terms of delayed ripening index. Inddition, the lower TSS/TA ratio in treated berries was due toower accumulation of TSS and especially to titratable acidityetention (Table 2).

.3. Nutritive parameters

The major sugars found in ‘Royal Autumn’ tablerapes were glucose and fructose, and concentrations wereery similar in recently harvested berries: 8.23 ± 0.43 and.30 ± 0.46 g 100 g−1, respectively. During cold storage, theevels of both sugars increased in control berries, with thencrease being higher during SL periods (Fig. 1). The accu-ulation of these sugars was significantly reduced when

ugenol and thymol were added to the packages, with noignificant differences attributed to the natural compound orpplied dose. On the other hand, the concentration of sucrose

as much lower than glucose and fructose, with levels atarvest of 0.35 ± 0.02 g 100 g−1 showing a slight decreasever storage (≈0.20 g 100 g−1) without significant differ-nces between control and treated berries (data not shown).

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or each parameter, similar small letters in rows are not significantly differentt P < 0.05 from levels at harvest and during storage. For each column, similarapital letters are not significantly different at P < 0.05 among treatments.

With respect to organic acids, the major one was tar-aric acid with levels of 0.40 ± 0.02 g 100 g−1 at harvest. Aignificant reduction was obtained for control berries whileetention was shown for treated fruit (Fig. 2). The net reten-ion was more pronounced after the addition of thymol thanugenol. Moreover, the results showed that maintenance ofartaric acid was dose-dependent, since higher concentra-ions of this organic acid was observed when 150 �l wasdded inside the packages. The remaining organic acids iden-ified and quantified occurred at much lower concentrationst harvest (mg): oxalic acid (59.19 ± 3.21 mg 100 g−1), cit-ic acid (10.83 ± 0.47 mg 100 g−1), succinic acid (144.54 ±.51 mg 100 g−1) and fumaric acid (2.43 ± 0.11 mg 100 g−1).ll showed the same behaviour, a reduction during storageithout any significant differences between treated and con-

rol berries (data not shown).

.4. Functional properties

The levels of ascorbic acid at harvest were8.23 ± 3.35 mg 100 g−1, which decreased throughoutold and storage and SL. However, control berries sufferedhe greatest losses of ascorbic acid, which were significanturing the first 28 days of storage (Fig. 3). On the contrary,he addition of eugenol and especially thymol slowed thescorbic acid loss.

Total antioxidant activity (TAA) was measured inoth skin and pulp. TAA of the skin at harvest was90.16 ± 66.80 mg equiv. ascorbic acid 100 g−1, while inhe flesh the activity was four-fold lower (139.60 ±.00 mg equiv. ascorbic acid 100 g−1). However, changes inAA were similar for both skin and flesh with a prolongediminution with time in control fruit either during cold stor-

ge or SL (Fig. 4). In the skin, when eugenol was added toAP packages, an increase in TAA was observed, the effect

eing dose-dependent. The addition of thymol led to main-enance of TAA with slightly higher activity for 150 than

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D. Valero et al. / Postharvest Biology and Technology 41 (2006) 317–327 321

Fig. 1. Glucose and fructose changes in MAP-packaged berries without (control) and with thymol (75 or 150 �l) or eugenol (75 or 150 �l) during cold storageor shelf life (SL). Data are the means ± S.E. (n = 10).

Fig. 2. Tartaric acid changes in MAP-packaged berries without (control) and with thymol (75 or 150 �l) or eugenol (75 or 150 �l) during cold storage or shelflife (SL). Data are the means ± S.E. (n = 10).

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Fig. 3. Ascorbic acid changes in MAP-packaged berries without (control) and with thymol (75 or 150 �l) or eugenol (75 or 150 �l) during cold storage or shelflife (SL). Data are the means ± S.E. (n = 10).

Fig. 4. Total antioxidant activity (TAA) changes in MAP-packaged berries (skin and pulp) without (control) and with thymol (75 or 150 �l) or eugenol (75 or150 �l) during cold storage or shelf life (SL). Data are the means ± S.E. (n = 10).

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D. Valero et al. / Postharvest Biology and Technology 41 (2006) 317–327 323

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ig. 5. Total phenolics changes in MAP-packaged berries (skin and pulp) wold storage or shelf life (SL). Data are the means ± S.E. (n = 10).

5 �l dose. In the pulp, both eugenol and thymol inducedhigher TAA during storage, the effect being more pro-

ounced and dose-dependent for eugenol. It is interesting tooint out that transfer from cold to SL for each sampling dateid not induce a significant loss of TAA in treated berries,hile it diminished in control grapes, for both skin andulp.

The content of total phenolics was also quantified in thekin and pulp of the berries. As occurred with TAA, the lev-ls of total phenolics at harvest were much higher in thekin (362.84 ± 21.92 mg equiv. gallic acid 100 g−1) than inhe pulp (17.39 ± 1.23 mg equiv. gallic acid 100 g−1). Con-rol berries exhibited a significant loss of total phenolics eithern the skin or pulp during cold storage or SL (Fig. 5). In con-rast, treated grapes showed a reduced rate of loss of totalhenolics in the skin at 1 ◦C to levels that were maintaineduring SL, and increases in the pulp independently of thetorage temperature. The effect was clearly dose-dependent

ith eugenol treatment being the most effective.As noted above, ‘Autumn Royal’ is a dark-purple

lack cultivar and showed a high concentration of totalnthocyanins in the skin at harvest (390.02 ± 25.66 mg

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(control) and with thymol (75 or 150 �l) or eugenol (75 or 150 �l) during

quiv. cyanidin-3-glucoside 100 g−1). During cold storage,significant loss of these pigments was detected in con-

rol berries, which was accelerated after the SL periodsith the levels of anthocyanins halved at the end of the

xperiment (Fig. 6). In treated grapes, maintenance of totalnthocyanins was detected for the thymol treatment, whiledelay in the pigment loss was shown for eugenol-treated

erries.

.5. Microbial parameters

The incidence of berry decay was affected by treatment.hus, in control grapes the fungal development started after8 days at 1 ◦C + SL, with percentage levels increasing dur-ng storage. On the contrary, in treated clusters decay waselayed with time and significantly reduced. Thus, at thend of the experiment, occurrence of decay was over 50%n control grapes and significantly reduced by the addition

f eugenol and thymol (Fig. 7). In addition, the effect wasose-dependent, since the lower percentages of decay werebtained for the higher dose (150 �l). The increase in berryecay was accompanied by a significant accumulation of

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ig. 6. Total anthocyanins changes in MAP-packaged berries (skin) withoutorage or shelf life (SL). Data are the means ± S.E. (n = 10).

thylene, which showed a concentration of 1.36 ± 0.09 �l l−1

n control berries after 56 days at 1 ◦C + SL and ≈0.2 �l l−1 inreated grapes without significant differences due to appliedompound or dose.

At harvest, table grapes had 2.3 ± 0.15 and 2.9 ±.17 log CFU g−1 for total mesophilic aerobic and yeastnd mould counts, respectively. After 28 and 56 days ofold storage, the microbial populations of treated grapesere significantly reduced, the reduction being slightlyore effective for yeast and mould (1.70–2.40 log CFU g−1)

han for mesophilic aerobic (2.20–2.40 log CFU g−1) counts.

n the contrary, significant increases in both yeast andould (4.2 ± 0.32 log CFU g−1) and mesophilic aerobic

4.2 ± 0.18 log CFU g−1) populations were observed for con-rol berries after 56 days.

ig. 7. Decay percentage of MAP-packaged berries without (control) andith thymol (75 or 150 �l) or eugenol (75 or 150 �l) during cold storage or

helf life (SL). Data are the means ± S.E. (n = 15).

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rol) and with thymol (75 or 150 �l) or eugenol (75 or 150 �l) during cold

.6. Sensory parameters

The highest scores after 56 days of cold storage and SLt 20 ◦C for fruit and rachis aspect, firmness, and crunchi-ess were given to the treated berries compared with con-rols. An inverse tendency was found when sweetness anduiciness were tasted, since control berries had significantlyigher scores (Fig. 8). Judges perceived development of “off-avours” in ‘Autumn Royal’ grapes but the percentage was

nfluenced by treatment. Thus, for control berries, 70% ofhe panellists found bad aromas and “off-flavours”, while forhose treated with thymol only 10% of panellists found theccurrence of “off-flavours”, this percentage being 15% forugenol treatment.

. Discussion

Table grapes, as for many food products, are naturallyerishable, as can be seen by the loss of quality in controlruit due to weight loss, skin colour changes, acceleratedoftening and increased maturity index throughout storage.hese changes were also accompanied by a high occurrencef decay, which could not be totally controlled using MAP.ll these changes were significantly delayed in those berries

reated with eugenol or thymol. Since there were not sig-ificant differences in the atmosphere composition withinackages, the induced benefit in the fruit quality parametershould be attributed to the essential oils themselves.

The higher reduction of weight loss by eugenol or thy-ol would indicate a role of these compounds in lowering

he dehydration process, as has been observed to occur inherry (Serrano et al., 2005a) and in ‘Crimson Seedless’ tablerape (Valverde et al., 2005), although the intrinsic mecha-ism is still unknown. In table grapes, colour and texture are

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D. Valero et al. / Postharvest Biology and

Fig. 8. Scores for the sensory analysis of MAP-packaged berries without(5(

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control) and with thymol (75 or 150 �l) or eugenol (75 or 150 �l) after6 days of cold storage + 2 days at 20 ◦C (SL). Data are the means ± S.E.n = 50).

mportant parameters from the consumer point of view. Skinolour of treated table grapes showed lower variations in L*,* and b* coordinates with respect to those values at harvest,hile control berries exhibited significant colour changes.t the optimum harvest date, ‘Autumn Royal’ table grapeas a purple-black skin colour due to the presence of antho-yanin compounds, from which the anthocyanin malvidin--glucoside has been found as major component (Valero etl., unpublished data). It has been reported that the ripeningrocess of table grapes is correlated with the anthocyanin con-entration and profile (Cantos et al., 2002). Thus, the lowerolour changes could be related to delays in the ripening pro-ess.

Softening contributes to quality loss in reducing the shelfife, but the addition of eugenol or thymol resulted in main-enance of flesh firmness during cold storage or slightlyeduction during SL. This effect has been reported in sweetherry (Serrano et al., 2005a) although the mechanism is

till unclear. Since the main cell wall degrading-enzymesesponsible for table grape softening are �-galactosidase,olygalacturonase and pectinmethylesterase (Nunan et al.,998), some additive effect of these compounds with MAP

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Technology 41 (2006) 317–327 325

n the reduction of the activity of these enzymes could bexpected.

Several authors have reported that TSS, TA and the ratioSS/TA are other important quality factors determining tablerape acceptability by consumers (Crisosto and Crisosto,002; Sonego et al., 2002). In this work, consumers gen-rally prefer sweet grapes but with a certain level of acidity.he main sugars found in ‘Autumn Royal’ table grape werelucose and fructose (ca. 8% each) while sucrose occurredt much lower concentrations. The TA was mainly due to theresence of tartaric acid (≈0.4%) while oxalic, citric, succinicnd fumaric acids occurred at milligram concentrations. Inur work, control berries exhibited a higher accumulation ofugars (glucose and fructose) and a significant reduction ofA (tartaric acid) which accounted for the higher TSS/TAatio over storage. On the contrary, the combination of MAPith eugenol and thymol led to a lower increase in the TSS/TA

ation throughout storage, showing that eugenol or thymolnduce a slower physiological maturation in table grapes.

The association between diet and health has been recog-ised since antiquity, and the role of fruit consumptionn preventing the risk to develop several degenerative dis-ases has been reported (Halliwell, 1996). The responsibleompounds for these beneficial properties for human healthre those with antioxidant activity, including carotenoids,scorbic acid, flavonoids, and phenolic compounds such asnthocyanins (Tomas-Barberan and Espın, 2001; Kalt, 2005).Autumn Royal’ table grapes could be considered as a goodource of these health-promoting compounds, as reported inhis work. It is interesting to point out that in this cultivar,AA was five-fold higher in the skin than in the pulp, asas been reported in other grape cultivars (Bartolome et al.,004). TAA potential of ‘Autumn Royal’ grapes could beonsidered as medium-high compared to other cultivars, inhich berries with highly pigmented skin exhibited higherAA (Kotamballi et al., 2002) than those with less red colourntensity (Cho et al., 2004).

In this work, the loss of quality during storage was alsoccompanied by diminution of the functional properties.hus, in control grapes total phenolics (skin and pulp), ascor-ic acid (pulp) and total anthocyanins (skin) were drasticallyeduced either during cold storage or further SL, which coulde responsible for the reduction of TAA found in both skinnd pulp. In contrast, the addition of eugenol or thymol insidehe packages led to a delay in the loss of total phenolicsnd total anthocyanins in the skin and ascorbic acid in theulp. In this tissue, the treated grapes exhibited a signifi-ant accumulation in total phenolics. The higher proportionf total phenolics and maintenance of total anthocyaninsere correlated with increases in TAA of the pulp and in

he skin of eugenol-treated berries, while maintenance wasbserved in the thymol-treated skins. In fact, TAA of the skin

as highly correlated (r2 = 0.99) with total phenols and total

nthocyanins, the same correlation being observed betweenAA of the pulp and the content of total phenolics and ascor-ic acid. In other table grape cultivars, TAA has also been

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26 D. Valero et al. / Postharvest Bio

orrelated with phenolic compounds (Davalos et al., 2005)nd anthocyanins (Kallithraka et al., 2005), these polyphe-ols being postulated for use as natural antioxidants in foodsBonilla et al., 1999; Pazos et al., 2005).

Eugenol and thymol have been described to exhibit a nat-ral antioxidant activity (Lee et al., 2005; Sacchetti et al.,005), close to that of vitamin C (Kim and Lee, 2004) andainly due to the presence of hydroxyl groups in the ben-

ene ring (Shahidi et al., 1992). Thus, the combined use ofoth compounds together with MAP storage would lead toaintenance or increase in the TAA of ‘Autumn Royal’ table

rapes.Decay was drastically reduced in those berries treated

ith eugenol or thymol, this reduction being accompanied byiminutions in either mesophilic aerobic or yeast and mouldounts from the values at harvest in treated berries, whilencreases were detected in control grapes. These results con-rm the antimicrobial effects described for these compoundsAppendini and Hotchkiss, 2002).

In a recent review, the shelf life and safety improvementrom adding essential oils to perishable foods such as fish,eat and bakery products has been reported (Holley andatel, 2005), but there is almost no evidence on the usef essential oils or their components in combination withAP in terms of maintenance of overall quality (organolep-

ic, sensory, nutritive and functional) in fruit, apart from ourrevious work on sweet cherry and ‘Crimson’ table grapesSerrano et al., 2005a; Valverde et al., 2005). The mechanismf action of the essential oils is attributed to their hydropho-icity, which enables them to partition in the lipids of theell membrane and thus disturb its integrity and the inor-anic ion equilibria (Lambert et al., 2001; Bagamboula et al.,004). The presence of the phenolic ring may be necessaryor the antimicrobial activity of eugenol and thymol (Ulteet al., 2002). In future, the effectiveness of combinations ofwo or more of these compounds (with different functionalroups) should be investigated. In addition, the effect of theure compounds on preventing decay in artificially inoculatedrapes would give an idea of the more appropriate concen-rations, bearing in mind maintenance of table grape qualityarameters.

cknowledgements

This work has been co-funded by Spanish Ministry of Sci-nce and Technology through project AGL2003-03257/ALInd European Commission with FEDER funds.

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