Changes in nutraceutical quality of tomato under different organic substrates

Yield and nutraceutical quality of Sahel tomato cultivar under shade net was evaluated using different mixtures of organic substrates in Coahuila, northern Mexico. The treatments consisted of mixtures using sand (S), vermicompost (VC), solarized bovine manure (SB), agricultural soil (AS) and mineralized compost (MC). The treatments were: T1 (S:VC, 80:20), T2 (S:SB, 80:20), T3 (S:SB:AS, 80:15:05), T4 (S:VC:AS, 80:15:05), T5 (S:MC, 80:20), and a control treatment T6 (S, 100%) with Steiner solution. Yield results showed that the best organic treatments were T5 and T4 with 3.48 and 3.33 kg plant-1, respectively; while the highest yield was in control (T6) with 3.71 kg plant-1. The highest phenolic content in the organic treatment of fruits was 56.94 mg equivalent of gallic acid per 100 g in fresh weight for T5, while the treatment with chemical fertilization obtained 49.82. The greatest antioxidant capacity was obtained in T4, with 478.34 μM equivalent of Trolox/100 g fresh weight. Two colors were evaluated (yellow and red) corresponding to two ripeness phases. Lycopene content in tomatoes was in average 32% higher in the red colored fruits than in the yellow ones, with 3.12 and 2.24 mg lycopene 100 g pulp-1 values, respectively. For the red ones, treatment T5 showed the highest value (3.52) and in yellow ones T1 reported 2.35, while T2 presented the lowest values for both colors. Organic fertilizers based on S:VC can induce good amounts of lycopene in fruits of both colors, as well as improve phenols and antiOX.


Received on February 13, 2017; accepted on December 11, 2017
vary depending on the species, ripeness and environmental conditions during crop's development.Other researchers (Bunea et al., 2012;Omar et al., 2012) have reported that application of organic manure on crops increases fruit phenolic content and antioxidant capacity.These results may be due to a high content of humic acids (Gutiérrez-Miceli et al., 2007) and low N content in the organic fertilizers.
Lycopene constitutes over 95.4% of the total compounds in the red ripeness state and is responsible for its characteristic color (Helyes & Lugasi, 2006).Lycopene is also very important since it is one of the major antioxidants consumed by humans in a regular diet.
However, few studies have focused on how to increase the amount of this compound in fruits.According Luna-Guevara & Delgado-Alvarado (2014), lycopene content is affected by biotic or environmental factors, among these the mineral nutrition and the growth conditions are determinant.
In this research, the impact of different organic substrates on the phytochemical quality of tomato fruits was studied.

MATERIAL AND METHODS
The study was conducted under shade net conditions in Technological Institute of Torreon (ITT), Coahuila, México (103°22'33"W, 25°36'36" N).Climate of this region is dry desert with rainfall in summer and cool winters.Precipitation is 241.9 mm and annual mean temperature is 21.5°C, ranging from 33.7°C maximum and 7.5°C minimum.The used shade house consisted of a metallic structure, 25 m length, 12 m width, and 3.0 m high, covered with anti-aphids mesh (40 x 26 threads cm -2 , cal.0.009).The experiment was conducted in spring-summer 2014 period, using Saladette tomato type, Sahel cultivar (Roger Syngenta).Germination took place in polystyrene trays of 200 cavities, using Peat Moss as substrate.One seed per cavity was deposited, covering it with black plastic until seed germination, with seeding date of February 15, 2014.Sprinkler irrigation was used twice a day to keep the rooting substrate moist.Fertilization based on urea and superphosphate (12-60) was applied at a dose of 0.50 g L -1 ; application was made daily at the first irrigation, starting application when 90% emerged.When plants had an approximate height of 15 cm and five to six true leaves, they were transplanted placing one seedling per pot, and using 10 kg black polyethylene bags.
Irrigation water was used in treatments T1, T2, T3, T4 and T5, applying 1.0 L pot -1 day -1 ; in T6, 1.0 L pot -1 day -1 of nutrient solution.Plants were guided to a single stem and pruning consisted of the elimination of auxiliary buds, additionally tutors were placed on the single stem and guided with raffia to help plants stay in place and hold the fruit's weight.
The number of cuts made was nine and average bunches per treatment were as follows: 9 (T1), 6 (T2), 5 (T3), 8 (T4), 10 (T5) and 8 (T6).The measured variables, considering four fruits pot -1 cut -1 , were: yield (Y), the fruits were harvested taking into consideration its commercial size, for which a Shimadzu digital analytical scale was used, calculating it based on fruits' weight in each replication per treatment.Result was expressed in kilograms per plant (kg plant -1 ).Fruit length (polar diameter) and fruit width (equatorial diameter) of the four harvested tomatoes were measured, and classified according to the Mexican Official Standard NMX-FF-031 (SAGARPA-ASERCA, 2008); this was conducted for each treatment, using a digital Vernier.Soluble solids in the red fruits were measured using a Sper scientific 300001 refractometer.
Determination of AntiOX and phenolic in the red fruits was carried out considering its physiological maturity at grade 5 and according to classification of the Mexico Supreme Quality brand (SAGARPA-ASERCA, 2008).Total antioxidant capacity was determined in Trolox equivalent, antioxidant capacity by the 1,1 biphenyl, 2-picrilhidrazil (DPPH) radical method with some modifications made according to in vitro DPPH method (Molyneux, 2004).The absorbance of the solution was read, adjusting it to a 515 nm wave length.A standard curve was prepared with Trolox (Aldrich, St. Louis, Missouri, USA), and results were reported as equivalent antioxidant capacity in μM equivalent in Trolox per 100 g fresh weight (μM equivalent in Trolox /100 g FW).Analyses were made in triplicate.
Total phenolic compound content was measured based on modification of the Folin-Ciocalteau (Esparza et al., 2006) method.Phenolic content was calculated by a pattern curve using gallic acid (Sigma, St. Louis, Missouri, USA) and results were reported in equivalent gallic acid mg per 100 g fresh weight (mg equivalent GA/100 g FW).
Analyses were made in triplicate.
Fruits that showed red and yellow colors were selected and frozen at -40°C for lycopene analysis, and it was extracted using the modified protocol of Mayeaux et al. (2006).Fruit colors (yellow and red) were analyzed by a Minolta CR-400 tristimulus colorimeter.For lycopene quantification, an Agilent 2100 Series high performance liquid chromatograph (HPLC) was used, where a Supelco Discovery ® C 18 (5 cm x 4.6 mm and 5 µm) column was installed.To set the operating conditions of the chromatograph, the technique reported by Anguelova & Warthersen (2000) was taken as basis, with different injection speeds and concentrations of the solvents in the mobile phase.A Sigma brand lycopene standard was used, its real concentration was determined by the lycopene molar absorptivity coefficient in hexane.For each experimental unit, lycopene was extracted from tomato pulp and immediately injected to the HPLC.This procedure was conducted three times for each experimental unit.To determine the lycopene concentration in each case, the peak area of the sample was compared to the peak area of the standard; results were reported in mg lycopene values per 100 g pulp -1 .
The experimental design, used to evaluate the effect of treatments with four replicates each, was completely randomized.Data were analyzed statistically by analysis of variance and means and was separated by the Tukey (P≤0.05)test using software package SAS Version 9.2 (SAS, 2009).

RESULTS AND DISCUSSION
The evaluated substrates determined the quality components of tomato fruit, since statistical analysis presented significant differences (P≤0.05) between treatments for fruit quality.
The highest average fruit weight was found in the organic treatment T5 (390 g) containing mineral compost, T4 (360 g) containing vermicompost and soil, and T1 (270 g) containing only vermicompost, being statistically equivalent to fruit's weight of T6 Steiner solution control treatment (283 g).The lowest average fruit weights were in treatments T2 and T3 with 140 and 103 g, respectively.Both treatments were mixed with solarized bovine manure (Table 1).Yields found in the organic substrate treatments with average weight of 252.6 g, were above the data reported by Marquez et al. (2008), who, working with vermicompost plus sand and vermicompost plus pearlite treatments, obtained a weight of 238.4 g.
For the polar diameter in organic treatments, the mean values were between 6.4 and 5.3 cm, having the highest values in T5, which contained mineralized compost (S:MC, 80:20).The best organic treatment (T5) was 27.0% higher than the chemical one; it is worth mentioning that fruits of this treatment are considered medium size.The equatorial diameter did not show significant differences (Table 2).These results are equal to those found by Moreno et al. (2012), who stated that the polar diameter of tomatoes (Miramar and Romina genotypes) using a mixture of earthworm humus and sand was 5.9 to 6.1 cm.Regarding the equatorial diameter, no significant difference was found in this study in a 4 to 4.5 cm range.De la Cruz et al. (2010) also did not find significant differences when using the compost and earthworm humus mixture in different proportions (100:00; 75:25 and 50:50).
For soluble solids content, according to the means comparison, no significant differences were found between substrates and sand mixtures, or in the control treatments.However, in this study, two treatments (T4 and T6), presented quality fruits in terms of soluble solids (Table 2).According to De la Cruz-Lazaro et al. (2009), tomato for fresh use must have contents over 4.5°Brix.
According to Zhai et al. (2015), soluble solids increase in tomato as the salinity of the irrigation water increased.In this study, different results were observed since high values of soluble solids were not associated with high electrical conductivity in the organic substrates.
In the mean comparison test for yield per plant, significant statistical differences were found between treatments, highlighting, as expected, the control treatment fertilized with inorganic nutritive solution (T6) with 3.71 kg plant -1 average.This was statistically equal to treatments T5 (S:MC, 80:20) and T4 (S:VC:AS, 80:15:05) with values of 3.48 and 3.33 kg plant -1 , respectively.These yields were in average, 12% lower than the control (Figure 1).
On the other hand, the solarized manure treatments in their higher proportion (S:SB, 80:20) and added with soil (S:SB:AS, 80:15:05) presented less than half yields, with respect to the best organic treatment which contains mineralized compost T5 (S:MC, 80:20).These results are lower than those found by Ortega-Martinez et al. (2010) of 4 kg plant -1 , using sawdust compost substrates; by De la Cruz-Lazaro et al. (2009) with 4 kg plant -1 using sand plus vermicompost (50:50) prepared with bovine manure; and by De la Cruz et al. (2010), with 5.73 kg plant -1 using tezontle and vermicompost and 5.58 kg plant -1 using tezontle and bovine manure in a 65:35 proportion.These three authors used Sun 7705 cultivar.
Total phenolic content and the antioxidant capacity of organic tomatoes relied on the type of substrate (Tukey, P<0.05).Phenolic content of the tomato produced in this study was 36.0 to 56.94 mg equivalent of gallic acid per 100 g in fresh weight for T1 (SVC/80:20) and T5 (S:MC, 80:20), respectively.That is to say, it increased by 56% when mineralized compost was used, with respect to vermicompost.However, the Steiner solution (T6), vermicompost with agricultural soil (T4), and solarized manure plus soil (T3) treatments were statistically equal (Figure 2).Results obtained in this work were higher than those reported by Zapata et al. (2007), who mention a 17.38±4.40and 18.91±5.75mg gallic acid/100 g phenolic content in fresh tomato in ripeness and commercial phase.
For this matter, it is stated that some fruits and vegetables contain certain chemical compounds (phytochemical) like phenols, which are believed to be related to the antioxidant synthesis as response to the subjection of the plant to oxidation stress, so substances are accumulated as a defense method.Oliveira et al. (2013) state the fruits from organic agriculture have a higher stress level, which along with the limitation of the nitrogen available in soil, make fruits 40% smaller.On the other hand, vitamin C and phenol concentrations were 55% and 139% higher than in conventional cultivation of tomatoes.
In this study, the greatest antioxidant capacity was obtained in treatment 4 (S:VC:AS, 80:15:05) being statistically equal to treatment 1 but statistically different to other treatments.Values found varied from 122 (T2) to 480 (T4) µM equivalent in Trolox/100 g FW ranges (Figure 2).Sahlin et al. (2004) reported that the amount of antioxidant molecules in tomatoes can be affected by several factors such as geographic location and environmental factors during cultivation, harvest time, storage conditions and fruit ripeness.Besides, the plants' response to stressing environmental conditions, like in organic production systems, generate defense mechanisms including antioxidants production (Winter & Davis, 2006).
The fruits' color is an important feature to the consumer's eye.In case of tomato, lycopene is responsible for its red color.Bio-synthesis of its pigmentation is genetically controlled and is affected by the environment (Brandt et al., 2006).In this study, lycopene content of harvested tomatoes depended on the type of substrate, that is, ANOVA showed significant differences (P<0.05).For this variable, two colors were evaluated (yellow and red) corresponding to two ripeness phases.
According to observations, the lycopene content on fruits was in average, 32% higher in the red colored ones than in the yellow ones, with 3.12 and 2.24 mg lycopene values, 100 g pulp -1 , respectively.For red color, treatment T5 presented the highest value, followed by treatment T1, treatment T4 and treatment T3; control treatment (T6) presented a value of 2.62, 34% lower than the highest treatment (T5).
It is worth noting that in the two analyzed colors, the control treatment (T6) obtained the lowest lycopene values (Figure 3).The amount of pigment found in this work was lower compared to the values reported by Gaspar-Peralta et al. (2012), with 9.6 to 16.8 mg/100 g wet base and 4.83 to 14.1 mg wet base in peel, reported by George et al. (2004).Arias et al. (2000), mention that the lycopene concentration increases with the ripeness of tomatoes, when chloroplasts change to chromoplasts and lycopene synthesis increases  causing red coloring.Luna-Guevara & Delgado-Alvarado (2014), state that lycopene content in tomatoes depends mainly on genetic (vegetable material), environmental (mineral nutrients, soil conditions, growing periods, etc.) and ripeness factors.Oliveira et al. (2013), state that among all factors that seem effective in enhancing the concentrations of phytochemicals in fruits and vegetables, stress emerges as especially promising.
With sand plus soil and vermicompost (S:VC:AS, 80:15:05) mixture, greater phenolic contents were obtained in the mineralized compost (T5), solarized manure plus soil (T3) and were statistically equal to Steiner solution (T6) in a 50 to 56.9 mg equiv.GA/100 g FW tomato range.Antioxidant capacity was higher in treatment 4 (S:VC:AS, 80:15:05), where the value was 480 µM equivalent in Trolox/100 g FW.Lycopene content in tomatoes was in average 32% higher in the red colored fruits than in the yellow ones, with 3.12 and 2.24 mg lycopene 100 g pulp -1 values, respectively.For the red ones, treatment T5 showed the highest value (3.52) and in yellow T1 reported 2.35, while T2 presented the lowest values for both colors.Organic fertilizers based on SVC can induce good amounts of lycopene in fruits of both colors, as well as improve phenols and antioxidant capacity.

Figure 1 .
Figure 1.Yield of tomatoes produced in planters with organic substrates mixtures in shade net system.Different letters inside each column indicate significant statistical difference (Tukey; P≤0.05).Lines over bars represent standard error for each treatment's mean.S= sand; VC= vermicompost; SB= solarized bovine manure; AS= agricultural soil; MC= mineralized compost.Coahuila, Mexico, Technological Institute of Torreon, 2014.

Figure 2 .
Figure 2. Average phenols content and average antioxidant capacity in tomato fruits produced in planters with organic substrate mixtures in shade net conditions.Different letters inside each column indicate a significant statistical difference (Tukey; P≤0.05).Lines over bars represent standard error for each treatment's mean.S= sand; VC= vermicompost; SB= solarized bovine manure; AS= agricultural soil; MC= mineralized compost.Coahuila, Mexico, Technological Institute of Torreon, 2014.

Figure 3 .
Figure 3. Lycopene colors, yellow and red, in tomatoes produced in planters with organic substrates in shade net conditions.Different letters inside each column indicate a significant statistical difference (Tukey; P≤0.05).Lines over bars represent standard error for each treatment's mean.S= sand; VC= vermicompost; SB= solarized bovine manure; AS= agricultural soil; MC= mineralized compost.Coahuila, Mexico, Technological Institute of Torreon, 2014.

Table 1 .
Chemical composition (dry weight) of the material used during evaluation of organic substrates for production of tomato under shade net conditions.Coahuila, Mexico, Technological Institute of Torreon, 2014.
Analyses were performed at the laboratory of the farming cooperative of the Comarca Lagunera.VC= vermicompost; SB= solarized bovine manure; MC= mineralized compost; AS= agricultural soil.

Table 2 .
Comparison of the fruit's mean weight, soluble solids, equatorial and polar diameters of tomatoes produced in organic substrates under shade net conditions.Coahuila, Mexico, Technological Institute of Torreon, 2014.different letters inside each column indicate a significant statistical difference (Tukey; P≤0.05).CV= coefficient of variation.S= sand; VC= vermicompost; SB= solarized bovine manure; AS= agricultural soil; MC= mineralized compost.