**3. Results and Discussion**

In Table 1 is reported, for each vineyard, the effect of the treatment and time of storage on quality attributes. Data of both vineyards were enough in agreement, and as will be better explained with data discussion, main differences were due to the higher quality of grapes from 4-year old vines, which degraded much slower, in comparison to the 14-year old grapes, particularly when stored in air, or kept on the plant. Treatment and time of storage affected more parameters for grapes from 14-year-old vines, than in the case of grapes from the 4-year-old vines. For grapes from 14-year-old vines, treatment influenced firmness, weight loss, hue angle, chroma, titratable acidity, phenol content, antioxidant activity, acetaldehyde, ethanol, citric acid and all sensorial parameters (except for sweetness). The time of storage affected most of parameters except for weight loss, phenols, antioxidant activity, fumaric acid, and fizzy taste score. On the other side, for grapes of 4-year-old vines, hue angle, acidity, phenols, were not affected by treatment and much less parameters were affected by time of storage. Since interaction between treatment and time of storage, was often significant when treatment was significant, and mostly for sensorial score, the simple effect of treatment for each quality attribute was evaluated at each sampling time.

As for gas evolution within packages, CO2 concentration was reduced for both experiment to 10% after 20 days of storage and remained constant until the end storage, while O2 stayed up to the atmospheric level (18–20%). After 28 days of storage, the CO2 and O2 concentrations were approximately 10.5 and 20%, respectively.

In Figure 1 is shown the effect of treatment on firmness during the storage; in particular firmness of berries held in MAP remained practically unchanged until the end of storage for both experiments, when grapes hold on the PLANT showed higher firmness values, compared to grapes stored in air and in MAP. For grapes of 4-year-old vines, a singular increase of firmness values was observed on berries left on the plant for 28 days, suggesting a concentration of pectin and cellulose when there was less competition for nutrients between fruit. As for berry stored in AIR weight loss increased to 3.7 and 2.2% at the end of the storage, for grapes of 4- and 14-year-old vines, respectively.

Figure 2 shows the effect of treatment on cluster and stalk appearance scores during storage. Here the main difference due to the age of the plant can be observed. Although for 4-year-old vines, there was no difference over storage time for cluster appearance of grapes stored on the plant and in MAP, for grapes from 14-year-old vines, cluster score was best preserved in MAP up to 21 days of storage, with no difference after 28 days. In this case, grapes stored on the vine degraded much faster when hold on the plant or in the cold room, whereas the presence of CO2 was effective on delaying ethylene effects [29,30] for grapes from both vineyards.



<sup>1</sup> AIR and MAP treatment.

**Figure 1.** Effect of storage treatment on firmness of "Italia" table grape from a 4-year (**A**) and 14-year-old (**B**) vineyard during storage. At each sampling time different letters indicate mean values significantly different (*p* < 0.05 and Tukey test; ns: not significant).

**Figure 2.** Effect of treatment on stalk and cluster appearance scores of "Italia" table grapes from a 4-year (**A**) and 14-year-old (**B**) vineyard during storage. At each sampling time different letters indicate mean values significantly different (*p* < 0.05 and Tukey test; ns: not significant).

In terms of visual appearance, the main effect of the different storage treatment was observed on the stalk, which showed dehydration and discoloration, for grapes stored in AIR, while less differences were observed for the berries, particularly for those from 14-year-old vines. Grape berries were less influenced by water loss, since they are well protected with waxy layers.

Stalk appearance showed a severe deterioration for grapes stored in AIR, whereas appearance scores remained almost unchanged for PLANT and MAP stored grapes. The score for AIR samples reached the value of 1 at 28 days, certainly for the observed dehydration, which occurred despite the protected conditions (macroperforated bag and humidified water pad). On the other hand, the high score values registered for samples in MAP are certainly related to low levels of dehydration but also to the effect of high-CO2 atmospheres on slowing down chlorophyll degradation rate. The effect of atmospheres with 29 kPa CO2 and 1 kPa O2 on chlorophyll retention has been demonstrated by Pariasca et al. [29] on pea pods and by Cefola et al. [31] on broccoli raabs stored with 10% CO2. Also, Silva-Sanzana et al. [22] reported that modified-atmosphere packaging helped to maintain green color of the stalk for "Red Globe" grapes stored for 90 days at 0 ◦C, with no negative effect on the quality of the berries. Similar results were found in "Autumn seedless" table grapes [20], indicating that clusters stored in air showed extreme browning of the stalk while clusters stored in CA (5 kPa O2+ 15 kPa CO2) and MAP (15 kPa O2+ 10 kPa CO2) had a good visual appearance at 60 days of storage at 0 ◦C and after additional 7 days in air at 15 ◦C.

Berry resistance to detachment decreased over time for all treatments; grapes stored in AIR showed a lower resistance to berry detachment than grapes hold on the PLANT or in MAP for grapes from 4-year old vines (Figure 3), and up to 21 days of storage for grapes from 14-year-old vine, where at 28 days PLANT showed highest value (3.7 N) and both air and MAP the same value of about 2.4 N. These results can be in part attributed to the different degree of water loss of stalk suffered by samples of different

treatments. Dehydration stress, in fact stimulates ethylene production which in turn favors the formation of the abscission layer, which reduces the force required for berry detachment. Pariasca et al. [29] and Bailén [30], found that the CO2 inhibits the abscission layer in MAP samples, because of the known competition of CO2 with ethylene on binding sites. In this case, for grapes of 14-year-old vines CO2 could had prevented ethylene effects, up to 21 days, but at the end of the storage in both MAP and AIR samples senescence processes were not inhibited, and no differences in berry detachment were observed.

**Figure 3.** Effect of treatment on flavor and berry detachment over storage of "Italia" table grapes from a 4-year vineyard (**A**) and on flavor and fizzy taste on grapes from 14-year-old (**B**) vineyard. At each sampling time different letters indicate mean values significantly different (*p* < 0.05 and Tukey test; ns: not significant).

As for the difference in color observed particularly for grapes of the oldest vine, a slightly higher decrease of b\*, Chroma (also for the 4-year-old-vine), and Hue angle values was observed for grapes stored on the plant, which also showed a higher increase of the a\* value. This may be explained by a higher enzymatic activity for grapes stored on the PLANT, and on the same time by the reduction of photosynthetic activity. Nonetheless difference in color were very minimum and not perceived by eyes. Regarding chemical attributes: titratable acidity, phenols, citric acid, and sucrose were significantly affected by the treatment, showing the same trend for grapes from both vineyards (data for grapes of 14-year-old vines is shown in Figure 4).

Titratable acidity and citric acid content increased after 28 days of storage for grapes store on the PLANT, which presented a higher content than AIR, while intermediate values were observed for MAP. Probably, the reduction of the fruit load could have led to a stimulation of the vegetative activity of the plant, inducing the increase of the acidity in the fruit. No differences for the other individual organic acids were found; tartaric and malic acids were the most abundant acids, followed by citric and succinic. In addition, traces of fumaric acid were also detected, in agreement with what reported in the literature for other varieties [32,33].

**Figure 4.** Effect of storage treatment on titratable acidity (**A**), citric acid (**B**), sucrose (**C**) and phenols (**D**) of "Italia" table grapes from a 14-year-old vineyard during storage. At each sampling time different letters indicate mean values significantly different (*p* < 0.05 and Tukey test; ns: not significant).

On the other hand, sucrose content decreased during storage; after 28 days, grapes stored in AIR showed a lower content (4.8 g·kg−1) than other treatments (about 7 g ·kg−<sup>1</sup> for both PLANT and MAP). As for glucose, at 28 days of storage was higher in MAP than in AIR, confirming a lower metabolism in MAP grapes probably associated with the presence of CO2. These results are confirmed by the soluble solids content that report a slight decrease during the storage without significant difference between treatments.

For all treatments and in both vineyards, an increase of phenolic content was also observed (much lower for grapes stored in MAP) immediately after 8 days of storage and up 21 days, followed by a reduction at the end storage. In Figure 4, data refer to 14-year-old vines, where after 28 days, grapes stored in AIR and on PLANT showed a higher phenolic content (1.92 and 1.80 g·kg−1, respectively) than the grapes stored in MAP (0.81 g·kg−1), but the same trend was observed also in the youngest vineyard. Probably, these results are associated with the biosynthesis of phenols, which is inhibited during postharvest storage in fruit and vegetables treated with elevated CO2 concentrations [34].

Regarding sensorial attributes, changes in flavor and presence of fizzy taste over time, are shown in Figure 3. It is important to notice that samples stored in MAP at the end of storage received the lowest flavor score, possibly related to the accumulation of CO2 in the cell sap in the form of carbonic acid which is the cause of the increase of the fizzy taste score, reported in figure for grapes from 14-year-old vines. On the other hand, it is important to highlight that the fizzy taste can also be due to fermentative processes caused by excessive accumulation of carbon dioxide and oxygen depletion typical of MAP [23] and associated with the accumulation of volatile compounds (ethyl acetate and ethanol). The latter aspect is more critical since when due to carbonic acid; it disappear after a few hours, with the evaporation of CO2.

With regard to aroma compounds a total of 21 volatiles, including six aldehydes (3-methylbutanal, pentanal, Z-2-butenal, hexanal, E-2-hexenal), six alcohols (ethanol, 1-hexanol, Z-3-hexen-1-ol, E-3-hexen-1-ol, 2-methyl-3-buten-2-ol, 1-pentanol), one ester

(ethylacetate), four terpenes (D-limonene, cis-, and trans-linaloloxide, linalool), three ketones (3-penten-2-one, 1-penten-3-one, 6-methyl-5-hepten-2-one) one furan derivative (2-ethylfuran), one acid (acetic acid), were found in both grapes from 14-year old vines and 4-year old vines. Grapes from 14-year-old vines also showed the presence of E-2-butenal. For both grapes from 14-year-old vines and 4-year-old vines, most of the volatiles were not affected by the storage treatment except for ethyl acetate and ethanol accumulating in MAP and some typical compounds better preserved in plant. Particularly ethyl acetate and ethanol showed the same trend during the storage, with significant differences only at 28 days of storage, in which the highest concentrations were found in the treatment with MAP (Figure 5). In general, the increase in ethyl acetate and ethanol concentrations has been also reported in the headspace of apples and strawberries stored in high CO2 due to the fermentative metabolism [35,36]. Also in this trial, the ethanol and ethyl acetate could be used as indicators to determine the grade of degradation of table grapes.

**Figure 5.** Effect of the storage treatment on the contents of ethyl acetate (**A**), ethanol (**B**), hexanal (**C**) (E-2-hexenal showed a similar trend), 1-hexanol (**D**), in grapes from 14-year-old vines (grapes from 4-year-old vines showed a similar trend). At each sampling different letters indicate significantly different mean values (*p* < 0.05 and Tukey test; ns: not significant).

On the other side, for both grapes from 14-year-old vines and 4-year-old vines, grapes kept on PLANT showed the highest concentration of E-2-hexenal, hexanal, and 1-hexanol, at the end of the storage (Figure 5), showing that grapes on the vine better maintained some typical compounds. The contents of all the other compounds, including e.g., the linalool, which is known to give floral notes, were not significantly different among the treatments.

Therefore, we can affirm that the MA packaging showed some advantages during the first 21 days of storage, as showed by the sensory evaluation of firmness, aroma, cluster, and stalk appearance, while at the end of storage the MA treatment suffered an accelerated process of fermentation likely characterized by the high content of ethanol and ethyl acetate, inducing the perception of fizzy taste.

#### **4. Conclusions**

Results of this experiment demonstrated that holding "Italia" table grapes on the PLANT, allowed a good preservation of the quality attributes (phenolic content, and flavor) compared to harvested product. Some additional benefits on cluster appearance score could be obtained by using MAP conditions, particularly if the grapes come from old vines, being more perishable than fruit from younger vine. Most volatile compounds did not change their concentration with the storage treatment, but ethyl acetate ant ethanol increased in MAP stored grapes at 28 days of storage, suggesting the occurrence of fermentation

processes confirmed by a highest perception of fizzy taste in grape stored with MAP conditions. Nonetheless, 21 days are a very reasonable time for the commercialization of this product, considering that normally for packaged fresh produce a shelf-life of 7–12 days is normally accepted. Therefore, depending on the market and distribution needs and to the age of the vineyard, different storage strategies may be applied.

**Author Contributions:** Conceptualization, G.C., M.L.A. Methodology, formal analysis, and investigation, F.P., S.P., G.C., M.L.A. Data curation, writing—original draft preparation, F.P. Writing—review and editing, supervision, G.C., M.L.A., S.P. Funding acquisition, G.C., M.L.A. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research received no external funding.

**Acknowledgments:** We thank the company Azienda Agricola F.lli Carpentiere Srl (Barletta, BT, Italy) for its kind cooperation in hosting this research in their vineyards.

**Conflicts of Interest:** The authors declare no conflict of interest.
