*2.5. Firmness*

The method developed by Maftoonazad and Ramaswamy [15] was used by means of a CT3 texture analyzer (Brookfield, Harlow, United Kingdom) adapted with a 5-mm conical strut probe. For the measurement, the epicarp was removed from both sides of the equatorial region. Ten repetitions were made per treatment and the results are expressed in Newtons (N).

#### *2.6. pH and Total Soluble Solids*

The pH was measured using a pH meter pH 211 microprocessor (Hanna, Rhode Island, USA). For the analysis, 10 g of mesocarp was homogenized with 90 mL of distilled water. The total soluble solids (TSS) was determined with a Pallete PR-101 refractometer (Atago, Washington, USA) and expressed in ◦Brix [16].

#### *2.7. Determination of Polyphenol Oxidase Activity*

The activity of polyphenol oxidase (PPO) was determined as reported by Vargas-Ortiz et al. [17]. For each treatment, 5 g of mesocarp was homogenized for 1 min in 15 mL of a 50 mM phosphate buffer (pH 6.5). The mixture centrifuged at 12000 rpm for 30 min at 4 ◦C and the supernatant used as the enzyme extract. Five hundred μL of 20 mM catechol was added as the substrate with 900 μL of 50 mM phosphate buffer (pH 6.5), and 100 μL of the enzyme extract. For the blank, 500 μL of 10% trichloroacetic acid (TCA) was added. The mixture was incubated for 20 min at 25 ◦C, and the reaction was stopped with the addition of 500 μL of 10% TCA. The absorbance was read at 410 nm using a spectrophotometer Jenway 6715 UV–Vis (USA). The PPO activity was reported as the increase in absorbance after the reaction time for 100 μL of extract.

#### *2.8. Determination of Colour*

The external and internal colour was determined using a CM-508d colourimeter (Minolta, Japan) to evaluate the parameters L\* (lightness), a\* (green to red) and b\* (blue to yellow). Five measurements were made on each fruit, and 10 fruits were examined per treatment per day of analysis [15], for internal colour, fruits were cut longitudinally and the same measurements were made as for epicarp.

#### *2.9. Bioactive Compounds and Antioxidant Activity*

#### 2.9.1. Extraction of Bioactive Compounds

The bioactive compounds of fruit were extracted according Vargas-Ortiz et al. [18]. Five grams of mesocarp was placed in a centrifuge tube, and 15 mL of ethanol/water solution (1:1) was added. The mixture was homogenized for 1 min at 4 ◦C and then was centrifuged at 12,000 rpm for 5 min at 4 ◦C. The supernatant was used for the determination of bioactive compounds and antioxidant activity.

#### 2.9.2. Determination of Total Phenols

The total phenolic content was determined by using the Folin–Ciocalteu assay as described by Villa-Rodríguez et al. [19] with some modifications. One milliliter of the extract was mixed with 5 mL of diluted Folin–Ciocalteau reagen<sup>t</sup> (1:10). After 6 min, 4 mL of Na2CO3 (20%) was added to the mixture, left for 2 h at room temperature and the absorbance against the reagen<sup>t</sup> blank was determined at 760 nm with an UV-Visible spectrophotometer. Total phenolic content was expressed as gallic acid equivalents/100 g of fruit (wet base). All the assays were performed in triplicate.

#### 2.9.3. Determination of Total Flavonoids

Total flavonoid content was measured by the aluminium chloride colorimetric assay using the method of Villa-Rodríguez et al. [19], with some modifications. An aliquot (1 mL) of extracts or standard solutions of quercetin was added to 4 mL of deionized water in a 10 mL flask. To the flask, 300 μL 5% NaNO2 was added and after five minutes, 300 μL 10% AlCl3. After another five minutes, 2 mL 1M NaOH was added and the volume was made up to 10 mL with deionized water. A blank was prepared in the same manner by using distilled water. The solution was mixed and absorbance was measured against the blank at 415 nm. The total flavonoid content was expressed as mg quercetin equivalents/100 g of fruit (wet base).

#### 2.9.4. Determination of Antioxidant Activity by Inhibiting the DPPH Radical

A 2,2-diphenyl-1-picrylhydrazyl (DPPH) reagen<sup>t</sup> was used to illustrate compounds with antioxidant activity [20]. A solution of DPPH 6.5×10−<sup>5</sup> M in 80% methanol was prepared, maintaining stirring for 2 h in darkness. Then, 0.5 mL of sample was mixed with 2.5 mL of the DPPH solution, and the mixture was stirred. The absorbance of the mixture was immediately read at a wavelength of 515 nm. The mixture was left to react in darkness for 1 h. As a blank, 80% methanol was used. The results obtained are expressed in mg of ascorbic acid equivalents/100 g of fruit (wet base).

#### 2.9.5. Determination of Antioxidant Activity by Inhibiting the ABTS Radical

Determination of antioxidant activity by inhibiting the radical 2,2-Azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) was performed as described by Vargas-Ortiz et al. [18] with some modifications. A 10 mL solution of 7 mM ABTS was prepared and reacted with 10 mL of 2.45 mM K2S2O8. The mixture was stirred for 16 h in a container in complete darkness. Afterwards the absorbance was measured in a spectrophotometer at 734 nm. The absorbance was adjusted with 20% ethanol to obtain a value of 0.7 ± 0.1. Two hundred μL of the sample was added to 2 mL of ABTS solution and allowed to react for 6 min and absorbance was measured at 734 nm. The results are expressed in mg of ascorbic acid equivalents /100 g of fruit (wet base).

#### *2.10. Structural Evaluation of the Epicarp*

Visually homogeneous fruits were selected and the fruit epicarp was cut into 1 cm<sup>2</sup> fragments of the equatorial part. Subsequently, according to the methodology developed by Hernández-Rivero et al. [21], the dissected material was fixed for 24 h in a mixture of formaldehyde, glacial acetic acid, 96% ethanol and distilled water at a ratio of 100:50:50:350. After this time, the tissue was washed with distilled water for 15 min, dehydrated, and infiltrated in an automatic tissue processor TP1020 (Leica, Germany) for 1 h in each of the following solutions: six sequential ethanol solutions at 60%, 70%, 80%, 90%, 96% and 100% and xylene. Then, the tissue was processed in two changes of paraplast solution for 2.5 h in each. The vegetal tissue was embedded in paraplast and cut transversely in 10 slices in a rotary microtome model 820 (Leica, Germany). The slices were mounted on slides, spread on a thermal plate at 25 ◦C for 24 h and stained with safranin-fast green to show the changes in the lignification of the cell walls, with the greater lignification staining red. The observations were made in an optical microscope model CX31RBSF (Olympus, Japan).
