Seed Priming Based on Iodine and Selenium Influences the Nutraceutical Compounds in Tomato (Solanum lycopersicum L.) Crop

The use of trace elements in agriculture as a complement to crop fertilization programs is a practice that is gaining importance and relevance worldwide. Iodine and selenium perform essential functions in human health, related to the proper functioning of the thyroid gland, acting as antioxidants and antiproliferatives, and their limited intake through food consumption can cause malnutrition, reflected in the abnormal development and growth of humans. This research aimed to evaluate the nutraceutical quality of tomato (Solanum lycopersicum L.) in response to seed priming based on KIO3 (0, 100, 150, 200, 250 mg L−1) and Na2SeO3 (0, 0.5, 1, 2, 3 mg L−1), performed by interaction from a 52-factorial design and by independent factors in a 24-h imbibition time. The tomato crop was established under greenhouse conditions in 10-L polyethylene containers containing peat moss and perlite 1:1 (v/v). Regarding non-enzymatic antioxidant compounds, lycopene, β-carotene and flavonoid contents in tomato fruits significantly increased with KIO3 and Na2SeO3 treatments; however, vitamin C content was negatively affected. KIO3 increased the phenol and chlorophyll-a contents of leaves. In relation to enzymatic activity, KIO3 positively influenced GSH content and PAL activity in tomato fruits. KIO3 also positively influenced GSH content in leaves while negatively affecting PAL and APX activities. Na2SeO3 favored GSH content and GPX activity in tomato fruits and leaves. Na2SeO3 negatively affected the antioxidant capacity of hydrophilic compounds by ABTS in fruits and leaves and favored hydrophilic compounds by DPPH in leaves. Seed imbibition based on KIO3 and Na2SeO3 is a method that is implemented in the tomato crop and presents interesting aspects that favor the nutraceutical quality of tomato fruits, which may contribute to increasing the intake of these minerals in humans through tomato consumption.


Introduction
Seed imbibition is considered a water-based technique that allows controlled hydration of the seed to trigger pregerminative metabolism but does not allow radicle emergence [1]. Seed imbibition is a common and effective process used to improve nutritional quality. The germination process is affected by external factors such as temperature, relative humidity, and light conditions, which can help or inhibit germination in relation to the nutritional reserve within the seed. During the germination process, some seed reserves are degraded  3 ) at a concentration of 1000 ppm was prepared. A mass of 168.59 mg of KIO 3 was gauged to 100 mL with distilled water. Dilutions of 2.5, 3.75, 5, and 6.25 mL of the stock solution were gauged to 25 mL with distilled water to obtain the treatments of 100, 150, 200, and 250 mg L −1 , respectively.
A stock solution of sodium selenite (Na 2 SeO 3 ) at a concentration of 1000 ppm was prepared. A mass of 21.89 mg of Na 2 SeO 3 was gauged to 100 mL with distilled water. Dilutions of 12.5, 25, 50, and 75 µL of the stock solution were gauged to 25 mL with distilled water to obtain the treatments of 0.5, 1, 2, and 3 mg L −1 , respectively.

Sowing and Planting
After the seed priming process, seeds were sown in polystyrene trays with peat moss and perlite 1:1 (v/v) as substrates. Seedlings were planted 35 days after sowing in 10-L plastic containers with peat moss and perlite 1:1 (v/v) as substrate. Tomato plants were arranged in the greenhouse in a randomized complete block design with a factorial arrangement of two factors (KIO 3 and Na 2 SeO 3 ) and five levels (concentrations in mg L −1 ) ( Table 1). Fertilization consisted of Steiner-type nutrient solutions [18], diluted in drip irrigation. The same substrate and fertigation conditions were used in the control and KIO 3 and Na 2 SeO 3 treatments to avoid another variation factor affecting the performance of the treatment.

Sampling
Samples of the leaves and ripe fruits of tomato plants were obtained 120 days after planting. Leaf samples were collected from the leaf tissue of fully extended young leaves from 12 plants with four replications. Samples for biochemical analysis were collected from five ripe fruits per treatment, with a uniform color and size corresponding to stage six of ripening [19]. Samples were stored at −80 • C, lyophilized in a 2.5 L FreeZone Benchtop Free Dry System freeze-dryer (LABCONCO, Kansas, MO, USA), and ground to a fine powder.

Vitamin C
The vitamin C content was determined by the 2,6-dichlorophenolindophenol titration method [20]. Here, 20 g of fresh fruit tissue was macerated by using a mortar and pestle with 10 mL of hydrochloric acid (2%), and 100 mL of distilled water was added and filtered with sterile gauze A 10-mL aliquot was taken and titrated with 2,6-dichlorophenolindophenol until a pinkish color was obtained (Equation (1)). The results were expressed in mg per 100 g of fresh tissue (mg 100 g −1 FW).

Total Phenols
Total phenolic compounds were determined using the Folin-Ciocalteu method from the extraction with water:acetone [21], made by adding 50 µL of the extract, 200 µL folinciocalteu reagent, 500 µL Na 2 CO 3 , and 5 mL distilled water to a test tube. The mixture was vortexed for 30 s and placed in a water bath at 45 • C for 30 min. The absorbance of the Antioxidants 2023, 12, 1265 4 of 20 mixture was read at a 750 nm wavelength. The results were expressed in mg of gallic acid equivalents per gram of dry tissue (mg GAE g −1 DW).

Total Flavonoids
Flavonoids were determined by the Dowd method, adapted by Arvouet-Grand et al. [22]. It was made by adding 2 mL of the extract and 2 mL of AlCl 3 (2%) methanolic solution to a test tube. The mixture was allowed to react for 20 min in the dark. The absorbance of the mixture was read at a 415 nm wavelength. The results were expressed in mg of quercetin equivalents per 100 g of dry tissue (mg QE 100 g −1 DW).

Chlorophyll
Chlorophyll content was quantified using the method proposed by Munira et al. [23]. Here, 1 g of fresh plant material was homogenized, and then 5 mL acetone (90%) was added. Additionally, 10 mg of magnesium carbonate (to protect and stabilize chlorophylls) was added, 2 mL of the extract was centrifuged at 9408× g for 5 min at 4 • C, and the supernatant was extracted. Chlorophyll-a and chlorophyll-b were quantified by reading the absorbances at 663 and 645 nm wavelengths, respectively. The results were computed (Equations (2)-(4)) and expressed in µg per gram of fresh tissue (µg g −1 FW).

Lycopene and β-carotene
Lycopene and β-carotene contents were determined according to Nagata and Yamashita [24]. Here, 100 mg of lyophilized tissue was mixed with 2 mL of hexane:acetone (3:2) solution. An aliquot was taken from the supernatant and the absorbances at 453, 505, 645, and 663 nm wavelengths were read. The results were computed (Equations (5) and (6)) and expressed in mg per 100 g of dry tissue (mg 100 g −1 DW). Samples of leaves and ripe fruits of tomatoes were freeze-dried and macerated by using a mortar and pestle; 200 mg of dry tissue and 20 mg of polyvinyl pyrrolidone were added in a 2-mL centrifuge tube; 1.5 mL of phosphate buffer (0.1 M, pH 7-7.2) was added; and the mixture was subjected to sonication for 5 min, and then centrifuged in a Prism C2500 refrigerated microcentrifuge (Labnet International Inc., Edison, NJ, USA) at 14,700× g for 10 min at 4 • C. The supernatant was collected and filtered with a 0.45-mmdiameter nylon membrane. Finally, the supernatant was diluted with phosphate buffer (1:20). This dilution was used to analyze the absorbances of reduced glutathione (GSH), glutathione peroxidase (GPX), phenylalanine ammonium lyase (PAL), catalase (CAT), and ascorbate peroxidase (APX) in a GENESYS 10S UV-Vis Spectrum (Thermo Fisher Scientific, Inc., Waltham, MA, USA), as well as the antioxidant capacity of ABTS and DPPH radicals in a BK-EL10C Elisa microplate reader (BIOBASE, Jinan, Shandong, China) at the corresponding wavelengths.

Reduced Glutathione (GSH)
GSH quantification was performed by the spectrophotometric technique [25]. It was made by adding 0.48 mL of the extract, 2.2 mL of Na 2 HPO 4 (0.32 M), and 0.32 mL of DTNB (1 mM) to a test tube. The sample was allowed to react for 15 min at room temperature, GPX was determined using the Flohé and Günzler [26] method, adapted by Xue et al. [25]. It was performed by adding 0.2 mL of the extract, 0.5 mL of GSH (1 mM), and 0.2 mL of Na 2 HPO 4 (0.067 M) to a test tube. The sample was placed in a water bath at 25 • C for 5 min, and 0.2 mL of H 2 O 2 was added. The mixture was allowed to react for 10 min, and the reaction was stopped with 1 mL of trichloroacetic acid. The mixture was centrifuged in a C2500 refrigerated microcentrifuge (Labnet Prism ® ) at 846× g for 10 min at 4 • C. Then, 0.48 mL of the obtained mixture, 2.2 mL of Na 2 HPO 4 (0.32 M), and 0.32 mL of 5.5 dithionis-2 nitro benzoic acid dye (1 mM) were added to a test tube. The mixture absorbance was read at a 412 nm wavelength. The results were expressed in units per gram of total protein (U g −1 TP), where U is equal to mM of GSH equivalents per mL per minute of dry tissue (mM GSHE mL −1 min −1 DW). PAL was determined according to Sykłowska-Baranek et al. [27]. It was performed by adding 0.1 mL of the extract and 0.9 mL of L-phenylalanine (6 mM) to a test tube. The mixture was incubated in a water bath for 30 min at 40 • C, and 0.25 mL of HCl (5 N) was added. The sample was placed in an ice bath, and 5 mL of distilled water was added. The mixture absorbance was read at a 290 nm wavelength. The results were expressed in units per 100 g of total protein (U 100 g −1 TP), where U is equal to µmol of trans-cinnamic acid equivalents per mL per minute of dry tissue (µmol TCAE mL −1 min −1 DW).
2.3.5. Catalase (CAT) (QE 1.11.1.6) CAT was determined by the spectrophotometric method [28], which is based on the quantification of the H 2 O 2 oxidation rate by absorbance difference (T 0 − T 1 ). T 0 was determined by adding 0.1 mL of the extract, 0.4 mL of H 2 SO 4 (5%), and 1 mL of H 2 O 2 (100 mM) to a test tube. T 1 was computed by adding 0.1 mL of extract and 1 mL of H 2 O 2 (100 mM), stirring for 1 min, and adding 0.4 mL of H 2 SO 4 (5%). The mixture absorbance was read at a 270 nm wavelength. The results were expressed in units per gram of total protein (U g −1 TP), where U is equal to mM of H 2 O 2 equivalents spent per mL per minute of dry tissue (mM H 2 O 2 E mL −1 min −1 DW).
2.3.6. Ascorbate Peroxidase (APX) (EC 1.11.1.11) APX quantification was performed according to the Nakano and Asada [29] method, which is based on the quantification of the H 2 O 2 oxidation rate by absorbance difference (T 0 − T 1 ). T 0 was determined by adding 0.1 mL of the extract, 0.5 mL of ascorbate (10 mg L −1 ), 0.4 mL of H 2 SO 4 (5%) to stop the reaction, and 1 mL of H 2 O 2 (100 mM) to a test tube. T 1 was computed by adding 0.1 mL of extract and 1 mL of H 2 O 2 (100 mM) to a test tube, stirring for 1 min, and adding 0.4 mL of H 2 SO 4 (5%) to stop the reaction. The mixture absorbance was read at a 266 nm wavelength. The results were expressed in units per gram of total protein (U g −1 TP), where U is equal to µmol of ascorbate oxidized equivalents per mL per minute of dry tissue (µmol AOE mL −1 min −1 DW).

Antioxidant Capacity of Hydrophilic and Lipophilic Compounds by ABTS
Antioxidant activity by the ABTS radical (2,2 -azino-bis-3-ethylbenzothiazolin-6-sulfonic acid) was determined by the spectrophotometric method [30]. It is based on the discoloration of the ABTS radical cation. This radical was obtained from the reaction of ABTS (7 mM) with potassium persulfate (2.45 mM) (1:1) under dark conditions at 25 • C for 16 h. Subsequently, it was diluted with ethanol (20%) to obtain an absorbance of 0.7 ± 0.01 at a 750 nm wavelength.
The antioxidant capacity of hydrophilic compounds by ABTS was determined by sample extraction performed with phosphate buffer, where 5 µL of extract and 245 µL of the dilution of ABTS radicals (7 mM) were placed in a microplate and allowed to react for 7 min in the dark. The absorbance of the extract was measured at a 750 nm wavelength.
The antioxidant capacity of lipophilic compounds as determined by ABTS was calculated by sample extraction performed with a hexane:acetone solution (3:2). Both the antioxidant capacity results of hydrophilic and lipophilic compounds by ABTS were expressed in µmol of Trolox equivalents per gram of dry tissue (µmol TE g −1 DW).

Antioxidant Capacity of Hydrophilic Compounds by DPPH
Antioxidant capacity by DPPH radical (2,2-Diphenyl-1-picrylhydrazyl) was performed according to Brand-Williams et al. [31]. The stock solution was prepared by mixing 2.5 mg of the DPPH radical with 100 mL of methanol. The absorbance of the solution was adjusted to 0.7 ± 0.02 at a 515 nm wavelength.
The antioxidant capacity of hydrophilic compounds by DPPH was determined as follows: 6 µL of the obtained extract was taken with phosphate buffer, and 234 µL of the DPPH radical was added. The decrease in absorbance of the extract after 30 min was measured at a 515 nm wavelength. The antioxidant capacity results of hydrophilic compounds by DPPH were expressed in µmol of Trolox equivalents per gram of dry tissue (µmol TE g −1 DW).

Statistical Analyses
The results were analyzed by analysis of variance to determine the variables that presented a significant statistical difference (p ≤ 0.05) so that the variables with significant effects were submitted to comparison means tests by Tukey (p ≤ 0.05) using the statistical software InfoStat ® 2020e.

Non-Enzymatic Compounds in Tomato Fruits by KIO 3 and Na 2 SeO 3 Interactions
The lycopene and β-carotene contents of tomato fruits were significantly modified by KIO 3 and Na 2 SeO 3 interactions. The lycopene content increased 110.6% in the 200-2 mg L −1 interaction and decreased 77.3% in the 200-3 mg L −1 interaction in relation to the control treatment. The β-carotene content increased 157.1% in the 200-1 mg L −1 interaction and decreased 90.5% in the 0-0.5 mg L −1 interaction in relation to the control treatment (0-0 mg L −1 interaction).
Phenol and flavonoid contents in tomato fruits significantly decreased in response to KIO 3 and Na 2 SeO 3 interactions. The phenol content increased by 9.7% in the 200-1 mg L −1 interaction and decreased by 32.3% in the 150-1 mg L −1 interaction in relation to the control treatment. Flavonoid content increased by 13.3% in the 250-1 mg L −1 interaction and decreased by 35% in the 200-3 mg L −1 interaction in relation to the control treatment.
The vitamin C content in tomato fruits was non-significantly influenced by KIO 3 and Na 2 SeO 3 interactions (Table 2).

Non-Enzymatic Compounds in Tomato Fruits by KIO 3 and Na 2 SeO 3 Factors
Regarding the potassium iodate factor in the tomato fruits, the vitamin C content significantly decreased by 5.2% with KIO 3 in the 150 mg L −1 treatment in relation to the control treatment. Phenol and flavonoid contents were not influenced by KIO 3 . Lycopene and β-carotene contents significantly increased with all KIO 3 treatments, where the 150 mg L −1 dose allowed reaching increments of 105 and 100% on lycopene and β-carotene contents, respectively, in relation to the control treatment, which contrasts with the results of vitamin C (Figure 1a).
Regarding the sodium selenite factor in the tomato fruits, the vitamin C content significantly increased by 9% with Na 2 SeO 3 in the 1 mg L −1 treatment in relation to the 0.5 mg L −1 treatment. The phenol content was not influenced by Na 2 SeO 3 . Flavonoid content increased by 8.6% with Na 2 SeO 3 in the 2 mg L −1 treatment and decreased by 7.2% in the 3 mg L −1 doses in relation to the control treatment. The lycopene content increased by 25% with Na 2 SeO 3 in the 0.5 and 2 mg L −1 treatments and decreased by 35% in the 3 mg L −1 treatment, in relation to the control treatment. The β-carotene content increased with Na 2 SeO 3 at 1-3 mg L −1 , reaching the highest increase by 72.7% in the 2 mg L −1 treatment in relation to the control treatment. The Na 2 SeO 3 treatment at 2 mg L −1 presented the best performance because it resulted in the highest increases in lycopene, β-carotene, and flavonoid contents in tomato fruits in relation to the corresponding control treatment ( Figure 1b).

Non-Enzymatic Compounds in Tomato Leaves by KIO3 and Na2SeO3 Interactions
The phenol content was significantly influenced by KIO3 and Na2SeO3 interactions, that is, increased by 31.6% in the 100-0.5 mg L −1 interaction and decreased by 28.4% in the 0-0.5 mg L −1 interaction in relation to the control treatment (0-0 mg L −1 interaction). The flavonoid content significantly decreased by 21.4% in the 100-3 mg L −1 interaction in relation to the control treatment. Chlorophyll and β-carotene were non-significantly influenced by KIO3 and Na2SeO3 interactions in relation to the respective control treatments.
Chlorophyll-a and total chlorophyll significantly increased by 3.2% and 2.4%, respectively, in the 100-0 mg L −1 interaction in relation to the 0-3 mg L −1 interaction, that is, removing the KIO3 and raising the Na2SeO3 doses (Table 3). Table 3. Effect of seed priming based on KIO3 and Na2SeO3 interactions on the non-enzymatic antioxidant compounds in tomato leaves.

Non-Enzymatic Compounds in Tomato Leaves by KIO 3 and Na 2 SeO 3 Interactions
The phenol content was significantly influenced by KIO 3 and Na 2 SeO 3 interactions, that is, increased by 31.6% in the 100-0.5 mg L −1 interaction and decreased by 28.4% in the 0-0.5 mg L −1 interaction in relation to the control treatment (0-0 mg L −1 interaction). The flavonoid content significantly decreased by 21.4% in the 100-3 mg L −1 interaction in relation to the control treatment. Chlorophyll and β-carotene were non-significantly influenced by KIO 3 and Na 2 SeO 3 interactions in relation to the respective control treatments.
Chlorophyll-a and total chlorophyll significantly increased by 3.2% and 2.4%, respectively, in the 100-0 mg L −1 interaction in relation to the 0-3 mg L −1 interaction, that is, removing the KIO 3 and raising the Na 2 SeO 3 doses (Table 3). Table 3. Effect of seed priming based on KIO 3 and Na 2 SeO 3 interactions on the non-enzymatic antioxidant compounds in tomato leaves.

Non-Enzymatic Compounds in Tomato Leaves by KIO 3 and Na 2 SeO 3 Factors
Regarding the potassium iodate factor in the tomato leaves, the phenol content significantly increased with all KIO 3 treatments, where the 100 and 200 mg L −1 treatments reached increments of 16.9% in relation to the control treatment. The chlorophyll-a content increased by 0.9% with KIO 3 in the 100 mg L −1 treatment in relation to the control treatment. Flavonoid, chlorophyll-b, and β-carotene contents were not influenced by KIO 3 (Figure 2a).

Non-Enzymatic Compounds in Tomato Leaves by KIO3 and Na2SeO3 Factors
Regarding the potassium iodate factor in the tomato leaves, the phenol content significantly increased with all KIO3 treatments, where the 100 and 200 mg L −1 treatments reached increments of 16.9% in relation to the control treatment. The chlorophyll-a content increased by 0.9% with KIO3 in the 100 mg L −1 treatment in relation to the control treatment. Flavonoid, chlorophyll-b, and β-carotene contents were not influenced by KIO3 (Figure 2a).
Regarding the sodium selenite factor in the tomato leaves, all the non-enzymatic antioxidant compounds were not significantly influenced by Na2SeO3 (Figure 2b). Regarding the sodium selenite factor in the tomato leaves, all the non-enzymatic antioxidant compounds were not significantly influenced by Na 2 SeO 3 (Figure 2b).

Enzymatic Activity in Tomato Fruits by KIO 3 and Na 2 SeO 3 Interactions
Regarding the tomato fruits, the (GSH) content and the PAL enzymatic activity were significantly increased by KIO 3 and Na 2 SeO 3 interactions: GSH by 35% in the 150-0.5 mg L −1 interaction and PAL by 441.7% in the 100-1 mg L −1 interaction in relation to the respective control treatments (0-0 mg L −1 interaction).
On the other hand, the enzymatic activities of GPX, CAT, and APX were not significantly influenced by KIO 3 (Table 4).

Enzymatic Activity in Tomato Fruits by KIO 3 and Na 2 SeO 3 Factors
Regarding the potassium iodate factor in the tomato fruits, the KIO 3 treatments at 150 and 200 mg L −1 significantly increased the GSH content by 9.5 and 14.3%, and the PAL enzymatic activity by 72 and 64%, respectively, in relation to the corresponding control treatments. GPX, CAT, and APX enzymatic activities were not significantly influenced by KIO 3 (Figure 3a).
Regarding the sodium selenite factor in the tomato fruits, the GSH content was significantly increased by Na2SeO3 by 14.3% and 9.5% in the 1 and 2-3 mg L −1 treatments, respectively, in relation to the corresponding control treatments. GPX enzymatic activity significantly increased by 64.3% by Na2SeO3 in the 3 mg L −1 treatment in relation to the control treatment ( Figure S2). PAL, CAT, and APX enzymatic activities were not significantly influenced by Na2SeO3 in relation to the respective control treatments. APX enzymatic activity significantly increased by 33.3% by Na2SeO3 in the 2 mg L −1 treatment in relation to the 1 mg L −1 treatment (Figure 3b).

Enzymatic Activity in Tomato Leaves by KIO3 and Na2SeO3 Interactions
GSH content and enzymatic activity of GPX in tomato leaves were significantly increased by KIO3 and Na2SeO3 interactions; GSH by 27.3% in the 200-0 mg L −1 interaction and GPX by 80% in the 150-0.5 mg L −1 interaction, in relation to the respective control treatments (0-0 mg L −1 interaction). The enzymatic activities of PAL, CAT, and APX were not significantly modified by KIO3 and Na2SeO3 interactions.
Higher enzymatic activities occurred for PAL in the 0-0.5 mg L −1 interaction, for CAT in the 250-0.5 mg L −1 interaction, and for APX in the 0-2 mg L −1 interaction (Table 5).  Regarding the sodium selenite factor in the tomato fruits, the GSH content was significantly increased by Na 2 SeO 3 by 14.3% and 9.5% in the 1 and 2-3 mg L −1 treatments, respectively, in relation to the corresponding control treatments. GPX enzymatic activity significantly increased by 64.3% by Na 2 SeO 3 in the 3 mg L −1 treatment in relation to the control treatment ( Figure S2). PAL, CAT, and APX enzymatic activities were not significantly influenced by Na 2 SeO 3 in relation to the respective control treatments. APX enzymatic activity significantly increased by 33.3% by Na 2 SeO 3 in the 2 mg L −1 treatment in relation to the 1 mg L −1 treatment (Figure 3b).

Enzymatic Activity in Tomato Leaves by KIO 3 and Na 2 SeO 3 Interactions
GSH content and enzymatic activity of GPX in tomato leaves were significantly increased by KIO 3 and Na 2 SeO 3 interactions; GSH by 27.3% in the 200-0 mg L −1 interaction and GPX by 80% in the 150-0.5 mg L −1 interaction, in relation to the respective control treatments (0-0 mg L −1 interaction). The enzymatic activities of PAL, CAT, and APX were not significantly modified by KIO 3 and Na 2 SeO 3 interactions.
Higher enzymatic activities occurred for PAL in the 0-0.5 mg L −1 interaction, for CAT in the 250-0.5 mg L −1 interaction, and for APX in the 0-2 mg L −1 interaction (Table 5).

Enzymatic Activity in Tomato Leaves by KIO 3 and Na 2 SeO 3 Factors
Regarding the potassium iodate factor in the tomato leaves, by increasing the KIO 3 concentration from 100 to 200 mg L −1 , the GSH content significantly increased from 10 to 20%, the enzymatic activity of PAL decreased from 23.9 to 28.9%, and the enzymatic activity of APX decreased from 51.4 to 37.8%. The enzymatic activities of CAT and APX were not significantly influenced by KIO 3 (Figure 4a).
Regarding the sodium selenite factor in the tomato leaves, the GSH content significantly decreased by Na 2 SeO 3 by 16.7 and 8.3% in the 0.5 and 2 mg L −1 treatments, respectively, in relation to the control treatment. The enzymatic activity of GPX significantly increased by 55.6% with Na 2 SeO 3 in the 0.5 mg L −1 treatment in relation to the control treatment. The enzymatic activities of PAL, CAT, and APX were not significantly influenced by Na 2 SeO 3 (Figure 4b).

Antioxidant Capacity in Tomato Fruits and Leaves by KIO3 and Na2SeO3 Interactions
Regarding the tomato fruits, the antioxidant capacity of hydrophilic compounds by the ABTS radical significantly decreased by 64.1% in the 200-2 mg L −1 (KIO3-Na2SeO3) interaction in relation to the control treatment (0-0 mg L −1 interaction). The antioxidant capacity of lipophilic compounds by ABTS and hydrophilic compounds by DPPH radicals was not significantly influenced by KIO3 and Na2SeO3 interactions. Higher values of antioxidant capacity of hydrophilic compounds by DPPH occurred in the 0-0.5 mg L −1 interaction (Table 6).
Regarding the tomato leaves, the antioxidant capacity of hydrophilic compounds by DPPH radical significantly increased by 187% in the 200-2 mg L −1 interaction, in relation to the control treatment. The antioxidant capacity of hydrophilic and lipophilic compounds by the ABTS radicals was not significantly influenced by KIO3 and Na2SeO3 interactions. Higher values of antioxidant capacity of hydrophilic compounds by ABTS occurred in the 150-3 mg L −1 interaction and for lipophilic compounds by ABTS in the 200-0.5 mg L −1 interaction (Table 6).

Antioxidant Capacity in Tomato Fruits and Leaves by KIO 3 and Na 2 SeO 3 Interactions
Regarding the tomato fruits, the antioxidant capacity of hydrophilic compounds by the ABTS radical significantly decreased by 64.1% in the 200-2 mg L −1 (KIO 3 -Na 2 SeO 3 ) interaction in relation to the control treatment (0-0 mg L −1 interaction). The antioxidant capacity of lipophilic compounds by ABTS and hydrophilic compounds by DPPH radicals was not significantly influenced by KIO 3 and Na 2 SeO 3 interactions. Higher values of antioxidant capacity of hydrophilic compounds by DPPH occurred in the 0-0.5 mg L −1 interaction (Table 6). Regarding the tomato leaves, the antioxidant capacity of hydrophilic compounds by DPPH radical significantly increased by 187% in the 200-2 mg L −1 interaction, in relation to the control treatment. The antioxidant capacity of hydrophilic and lipophilic compounds by the ABTS radicals was not significantly influenced by KIO 3 and Na 2 SeO 3 interactions. Higher values of antioxidant capacity of hydrophilic compounds by ABTS occurred in the 150-3 mg L −1 interaction and for lipophilic compounds by ABTS in the 200-0.5 mg L −1 interaction (Table 6).

Antioxidant Capacity in Tomato Fruits and Leaves by KIO 3 and Na 2 SeO 3 Factors
Regarding the potassium iodate factor, the antioxidant capacity of hydrophilic compounds by ABTS and by DPPH and of lipophilic compounds by ABTS was not significantly influenced by KIO 3 in tomato fruits ( Figure 5a) and leaves (Figure 6a). The antioxidant capacity of lipophilic compounds by ABTS in tomato fruits increased 88.6% with KIO 3 in the 100 mg L −1 treatment in relation to the 200 mg L −1 treatment (Figure 5a).
Regarding the sodium selenite factor in the tomato fruits, the antioxidant capacity of hydrophilic compounds by ABTS and by DPPH was negatively affected by Na 2 SeO 3 , where the 2 mg L −1 treatment presented higher inhibition of these parameters by 46.9 and 37.7%, respectively (Figure 5b).
Regarding the sodium selenite factor in the tomato leaves, the antioxidant capacity of hydrophilic compounds by DPPH significantly increased by Na 2 SeO 3 35.4%, 71.8%, and 54.6% in the 1, 2, and 3 mg L −1 treatments, respectively. The antioxidant capacity of hydrophilic compounds by ABTS significantly decreased by 15.9% by Na 2 SeO 3 in the 2 mg L −1 treatment in relation to the control treatment. The antioxidant capacity of lipophilic compounds by ABTS was not significantly influenced by Na 2 SeO 3 in tomato fruits and leaves (Figure 6b).

Discussion
Implementation of techniques such as seed imbibition is an effective method to improve the response of plants to biotic and abiotic stress conditions (Table S1) through the alteration of antioxidant metabolism [10]. Important findings of seed imbibition are reported, such as the antioxidant response of broccoli influenced by selenium [32], the salt stress tolerance of strawberries influenced by iodine species [33], and the functional effects of selenium in crucifers [34].
Plants can absorb different chemical elements from the soil or the nutrient solution, whether these elements are beneficial or toxic [35]. Selenium can be absorbed through the roots in the form of selenate (SeO4 2− ), selenite (SeO3 2− ), and organic Se compounds such as selenocysteine (SeCys) and selenomethionine (SeMet) ( Table S2), but selenides or elemental Se cannot be absorbed [36]. The use of selenium in plants has been reported to have positive effects on glutathione content, because selenium increases sulfur (S) receptors and consequently increases the absorption of both elements, favoring the synthesis

Discussion
Implementation of techniques such as seed imbibition is an effective method to improve the response of plants to biotic and abiotic stress conditions (Table S1) through the alteration of antioxidant metabolism [10]. Important findings of seed imbibition are reported, such as the antioxidant response of broccoli influenced by selenium [32], the salt stress tolerance of strawberries influenced by iodine species [33], and the functional effects of selenium in crucifers [34].
Plants can absorb different chemical elements from the soil or the nutrient solution, whether these elements are beneficial or toxic [35]. Selenium can be absorbed through the roots in the form of selenate (SeO4 2− ), selenite (SeO3 2− ), and organic Se compounds such as selenocysteine (SeCys) and selenomethionine (SeMet) ( Table S2), but selenides or elemental Se cannot be absorbed [36]. The use of selenium in plants has been reported to have positive effects on glutathione content, because selenium increases sulfur (S) receptors and consequently increases the absorption of both elements, favoring the synthesis

Discussion
Implementation of techniques such as seed imbibition is an effective method to improve the response of plants to biotic and abiotic stress conditions (Table S1) through the alteration of antioxidant metabolism [10]. Important findings of seed imbibition are reported, such as the antioxidant response of broccoli influenced by selenium [32], the salt stress tolerance of strawberries influenced by iodine species [33], and the functional effects of selenium in crucifers [34].
Plants can absorb different chemical elements from the soil or the nutrient solution, whether these elements are beneficial or toxic [35]. Selenium can be absorbed through the roots in the form of selenate (SeO 4 2− ), selenite (SeO 3 2− ), and organic Se compounds such as selenocysteine (SeCys) and selenomethionine (SeMet) ( Table S2), but selenides or elemental Se cannot be absorbed [36]. The use of selenium in plants has been reported to have positive effects on glutathione content, because selenium increases sulfur (S) receptors and consequently increases the absorption of both elements, favoring the synthesis of secondary metabolites [37].
Plant cells produce free oxygen and its derivatives, such as reactive oxygen species (ROS), which are used as signaling molecules in plants in signal translation in response to environmental conditions; this triggers antioxidant defense mechanisms. In this context, it has been shown that the optimal addition of iodine ( Figure S3) and selenium presents an alteration in the ROS production system [38,39], where the enzymatic defense systems, such as catalase, peroxidase, superoxide dismutase, glutathione peroxidase, and ascorbate peroxidase, and non-enzymatic antioxidants, such as glutathione, ascorbate, tocopherols, and phenolic compounds, are activated ( Figure S4) to reduce the excessive ROS production [40]. For its part, the SOD enzyme dismutases the O 2 − into hydrogen peroxide H 2 O 2 and molecular oxygen O 2 , and later the catalase (CAT) degrades the H 2 O 2 into oxygen and water, while the ascorbate peroxidase uses the ascorbic acid as a donor to stimulate the degradation of H 2 O 2 , and the reduced glutathione is responsible for the production of ascorbic acid [41]. Glutathione reductase catalyzes the regeneration of reduced glutathione (GSH) from glutathione disulfide (GSSG) with NADPH as the reducing agent. GSH eliminates H 2 O 2 by non-enzymatic reaction with O 2 − and OH − , likewise, GSH has the ability to replenish ascorbic acid through the ascorbate-glutathione cycle, which is of great importance for the antioxidant system [42].

Non-Enzymatic Compounds
In this research, the use of iodine and selenium concentrations applied to tomato crops by seed priming presented increases in the content of flavonoids, lycopene, and carotene in tomato fruits ( Figure S2). These results agree with those reported by Gaucin-Delgado et al. [43], who applied 2 mg L −1 Na 2 SeO 4 in the nutrient solution, presenting an increase in the content of phenols in the tomato crop. Likewise, Cunha et al. [44] and Ishtiaq et al. [45] indicated that the use of selenium presents an increase in chlorophyll, carotenes, and phenolic compounds when using concentrations of 7.5 and 15 µg kg −1 , while Sabatino et al. [46] indicated that the use of concentrations of 2 and 4 µmol of SeO 2 presented a higher content in carotenes in relation to the control. Phenol and vitamin C contents in tomato fruits did not present significant effects between treatments, which are similar to those reported by Smoleń et al. [47], who indicated that the use of KIO 3 in conjunction with Na 2 SeO 3 at concentrations of 30 and 8.5 µg dm −3 , respectively, did not present a significant effect in relation to the control.
The use of KIO 3 influenced an increase in the phenol and chlorophyll-a contents; however, the Na 2 SeO 3 treatments did not significantly modify these parameters in the leaves in relation to the control. Similar results were reported by Jerse et al. [48], who mentioned that the use of Na 2 SeO 3 at 10 mg L −1 in conjunction with KIO 3 at concentrations of 1000 mg L −1 did not present an effect on photosynthetic compounds, likewise indicating that the use of iodine concentrations and selenium separately reduced the dry matter content. However, when both elements interacted, there was a higher biomass content, which is similar to that reported by Smoleń et al. [49], who found that the separate use of selenium and vanadium promotes iodine uptake in plants.
Wang et al. [50] indicated that imbibition treatments present an increase in the content of polyphenols in the rye, which is attributed to the synthesis or activation of a variety of hydrolytic enzymes, causing different alterations in the structure or the synthesis of new compounds with high bioactivity and nutritional value. On the other hand, Vicas et al. [51] indicated that the use of selenium nanoparticles did not affect the phenol content of the broccoli crop. Likewise, Islam et al. [52] indicated that at a higher concentration of Na 2 SeO 3 , the phenol content begins to decrease. In the same way, Shohag et al. [53] indicated that the imbibed soybean and bean seeds presented a decrease in the phenol content in seeds and sprouts. This decrease in the phenol content is attributed to the imbibition time ( Figure S1) since it is considered that the longer the imbibition time, the greater the water absorption, which presents a dilution effect.
Jerše et al. [48] indicated that the use of iodine and selenium in the imbibition of seeds is a viable method because the enrichment of the pea shoots was achieved with the use of both elements; however, the uptake depends on the shape and/or combinations of the elements. The same effect was reported by Deng et al. [54] and Radawiec et al. [55], who indicated that in osmoconditioning treatments with iron, copper, manganese, zinc, selenium, and iodine in soybean and wheat seeds, they present an increase in the speed of germination and accumulation of these compounds in the shoots, for which they define seed imbibition treatments as a simple and highly efficient technique to increase the content of organic mineral elements in sprouts.

Enzymatic Activity
Regarding the enzymatic activity, the use of KIO 3 presented an improvement in the GSH content and a greater enzymatic activity in PAL ( Figure S3); similar results are reported by Blasco et al. [56], who indicated that the use of iodide (I − ) and iodate (IO 3 − ) in lettuce plants presents an increase in antioxidant enzymes. On the other hand, Na 2 SeO 3 presented a higher GSH and GPX content ( Figure S2); similar results were reported by Zhu et al. [13] and Rady et al. [57], where the use of selenium concentrations favors the increase in the GSH and GPX contents in the tomato crop. The increase in the GHS content is beneficial since high concentrations are needed to overcome oxidative stress in chloroplasts and other organelles [58].
Cao et al. [59], Diao et al. [60], and Kumar et al. [10] indicated that the use of Na 2 SeO 3 increases the enzymatic activity compared to the control, presenting an increase in GPX, CAT, and APX, while Nawaz et al. [61] indicated that the enzymatic activity of CAT and APX is increased in seed imbibition treatments with Se. Hu et al. [62] indicated that the use of selenium in the seed imbibition solution presents an increase in the α-amylase content, an increase in the sugar content, and an increase in the enzymatic activity of superoxide dismutase (SOD), glutathione peroxidase (GPX), and catalase (CAT), in addition to presenting a higher total chlorophyll content and Se content in the seedlings; however, this depends on the imbibition time of the seeds. Nawaz et al. [61] indicated that using selenium and exogenous zinc in seed's imbibition increases germination index, vigor, and enzymatic antioxidants such as catalase, guaiacol peroxidase, superoxide dismutase, and ascorbate peroxidase.

Antioxidant Capacity
Regarding the antioxidant capacity, there was a positive response when using Na 2 SeO 3 in the leaves by DPPH of hydrophilic compounds, while Fuentes et al. [63], Medrano Macías et al. [33], and Sarrou et al. [64] indicated that the use of KIO 3 does not affect the antioxidant capacity in strawberry and tomato crops.

Conclusions
The application of potassium iodate and sodium selenite in tomato crops by seed imbibition treatments influenced significant changes in non-enzymatic antioxidant compounds, such as phenols, lycopene, β-carotene, and reduced glutathione, as well as on enzymes, that is, phenylalanine ammonium lyase, catalase, and ascorbate peroxidase, both in tomato leaves and fruits; however, the same treatments influenced not significant changes in the antioxidant capacity. Seed priming based on trace elements is a useful and simple technique to perform in agricultural and horticultural production systems. Although this method does not present inconvenience due to the low concentrations of trace elements required, it is necessary to carry out more studies to establish the optimal concentrations according to the crop and the form of application, which allow for improvement of the desired indicators, such as the antioxidant compound pool of the edible organs of the plants, understand the balance and pathway of trace elements in the plant, and the benefits of biofortification of the fruits.

Supplementary Materials:
The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/antiox12061265/s1. Table S1: List of some patented seed priming treatments commercially available; Table S2: Summary of selenoprotein functions; Figure S1: Ranges of selenium application in several crops, applications form to achieve benefits such as biofortification and stimulation; Figure S2: Biochemical and ionomic effects of selenium and nanoselenium application in plants; Figure S3: Uptake, transport and metabolism of iodine in plants; Figure S4: Iodine as a modulator of antioxidants.