4.3.3. Weight Loss and Fruit Firmness

Fruit weight was recorded on the harvesting day (day 0) for each fresh produce (*n* = 6) per treatment and every other day, up to the last day of storage (day 14). Fruit firmness was measured at two points on the shoulder of each tomato fruit by applying a plunger of 3 mm in diameter at a speed of 2 mm s−<sup>1</sup> and the penetration depth was 12 mm, using a texture analyzer (TA.XT plus, Stable Micro Systems, Surrey, UK). The amount of force (in Newtons; N) required to break the radial pericarp (i.e., surface) of each commodity (*n* = 6) was recorded at ambient temperature (22–24 ◦C).

## 4.3.4. Soluble Solids, Titratable Acidity, Ascorbic Acid and Carotenoids

Total soluble solids (TSS) concentration was determined in triplicate from the juice obtained from two pooled fresh fruits for each replication (*n* = 3) with a temperaturecompensated digital refractometer (model Sper Scientific 300017, Scottsdale, AZ, USA) at 20 ◦C, and results were expressed in ◦Brix. Titratable acidity (TA) was measured via potentiometric titration (Mettler Toledo DL22, Columbus, OH, USA) of 5 mL of fruit juice diluted to 50 mL with distilled water, using 0.1 N NaOH up to pH 8.1. Results were expressed as g of citric acid per 1 L juice (g citric acid L−<sup>1</sup> juice). The fruit sweetness/ripening index was calculated using TSS/TA ratio.

Ascorbic acid (AA) was determined by the 2,6-Dichloroindophenol titrimetric method as described previously [47]. An aliquot of 5 mL of pooled tomato juice was diluted with 45 mL of oxalic acid 0.1% and was titrated by the dye solution until the color changed. Data were expressed as mg of AA per gram of fresh weight (mg AA g−<sup>1</sup> Fw).

Carotenoids (lycopene and *β*-carotene) for tomatoes were determined according to the method described by Nagata and Yamashita [48]. Briefly, 1 g of blended tomatoes was homogenized with 20 mL of acetone:hexane 4:6 (*v:v*) and after sonication and vigorous vortex the two phases were separated automatically. An aliquot from the upper phase was used for absorbance measurement at 663, 645, 505 and 453 nm in a spectrophotometer, using a reference of acetone:hexane (4:6) ratio. Lycopene and *β*-carotene contents were calculated according to the Nagata and Yamashita [48] equations:

Lycopene (mg 100 mL−<sup>1</sup> of extract) = <sup>−</sup>0.0458 <sup>×</sup> A663 + 0.204 <sup>×</sup> A645 + 0.372 <sup>×</sup> A505 <sup>−</sup> 0.0806 <sup>×</sup> A453. (1)

*<sup>β</sup>*-Carotene (mg 100 mL−<sup>1</sup> of extract) = 0.216 <sup>×</sup> A663 <sup>−</sup> 1.22 <sup>×</sup> A645 <sup>−</sup> 0.304 <sup>×</sup> A505 + 0.452 <sup>×</sup> A453. (2)

Results were expressed as mg per gram of fresh weight (mg g−<sup>1</sup> Fw).

#### 4.3.5. Total Phenolics and Antioxidant Activity

Total phenolic content was measured in blended fruit tissue (1 g) extracted with 10 mL of 50% (*v*/*v*) methanol, as reported previously [43]. Results were expressed as mg gallic acid equivalents (GAE) per gram of fresh weight (mg GAE g−<sup>1</sup> Fw). The antioxidant activity was determined using the ferric-reducing antioxidant power (FRAP) and 2,2-diphenyl-1 picrylhydrazyl (DPPH) radical-scavenging activity assays (at 593 and 517 nm, respectively) as described by Chrysargyris et al. [43]. The results were expressed in mg trolox per gram of fresh weight (mg trolox g−<sup>1</sup> Fw). All biological samples were analyzed in triplicate.

#### 4.3.6. Damage Index (Hydrogen Peroxide and Lipid Peroxidation)

Hydrogen peroxide (H2O2) levels were estimated using the procedure previously described by Loreto and Velikova [49]. After measuring the optical density (OD) at 390 nm, results were expressed as μmol of H2O2 per gram of fresh weight (μmol g−<sup>1</sup> Fw). Lipid peroxidation was determined with the 2-thiobarbituric acid reactive substances (TBARS) assay according to de Azevedo Neto et al. [50]. The absorbance was measured at 352 nm (discarding the non-specific absorbance at 600 nm) and results were expressed as nmol malondialdehyde (MDA) per gram of fresh weight (nmol g−<sup>1</sup> Fw).
