4.3.7. Enzymatic Antioxidant Activity

Fresh fruit tissue, powdered with liquid nitrogen (0.5 g) was homogenized with 4 mL of 50 mM potassium phosphate buffer (pH 7.0), which contained 1 mM phenylmethylsulfonyl fluoride (PMSF), 1 mM ethylenediaminetetraacetic acid (EDTA), 1% *w*/*v* polyvinylpolypyrrolidone (PVPP) and 0.05% Triton X-100. Samples were then centrifuged at 16000 g for 20 min, at 4 ◦C, and the supernatant was used for the determination of the antioxidant enzyme activity. Catalase (CAT, EC 1.11.1.6) activity was determined by following the consumption of H2O2 at 240 nm for 3 min. Results were expressed as CAT units per milligram of protein (units mg−<sup>1</sup> protein), using the extinction coefficient of 39.4 mM cm−<sup>1</sup> [51].

Superoxide dismutase (SOD, EC 1.15.1.1) was assayed as described by Chrysargyris et al. [52]. The reaction of 13 mM methionine, 75 μM nitro blue tetrazolium (NBT), 0.1 mM EDTA, 2 μM riboflavin and extract, was started after the addition of riboflavin. Reaction was then incubated under a light source of two 15-watt fluorescent lamps, for 15 min. The absorbance was determined at 560 nm and the activity was expressed as SOD units per mg of protein (units mg−<sup>1</sup> protein). One unit of SOD activity was defined as the amount of enzyme required to cause 50% inhibition of the NBT photoreduction rate.

Peroxidase activity (POD, EC 1.11.1.7) was determined according to the method used by Tarchoune et al. [53], using pyrogallol as substrate. The increase in absorbance at 430 nm was measured on a kinetic cycle for 3 min, and results were expressed as units of POD per milligram of protein (units mg−<sup>1</sup> protein).

An aliquot of the extract was used to determine the protein content by the Bradford method [54], with bovine serum albumin (BSA) as the protein standard.

#### 4.3.8. Sensory Evaluation

Fresh produce marketability, aroma and appearance were recorded by at least six panelists to compare the external visual aspect and marketability of treated and control fresh produce after 7 and 14 days of storage at 11 ◦C. Aroma was evaluated by using a 1–10 scale, with 1: bad aroma but not EP odor; 3: EP odor with some unpleasant smell; 5: EP smell but it is pleasant; 8: less tomato-like; 10: intense tomato-like. Appearance was evaluated by using a 1–10 scale, with 1: green color of 50%; 3: yellow-green; 5: orange; 8: red; 10: deep red. Marketability was evaluated by using a 1–10 scale, with 1: not marketable quality (i.e., malformation, wounds, infection); 3: low marketable with malformation; 5: marketable with few defects i.e., small size, decolorization (medium quality); 8: marketable (good quality); 10: marketable with no defects (extra quality).

#### *4.4. Statistical Analysis*

Statistical analysis was performed using IBM SPSS version 22 comparing data means (± standard error-SE) with one-way ANOVA, and Duncan's multiple range tests were calculated for the significant data at *p* = 0.05. Measurements were done in three (preliminary test) or three or six (main study) biological replications/treatment (each replication consisted of a pool of two to three individual measures/sample) on different assays.
