Urochloa Hybrid Submitted to Biostimulant Application in Grazing Simulation

The objective of this work was to evaluate the performance of Urochloa hybrid Convert HD364 applied in different doses as a vegetable biostimulant under grazing simulation in intermittent stocking in the city of Uberlândia, Minas Gerais. The treatments consisted of a control (absence of biostimulant) and in 5 doses of biostimulant, 0.25; 0.5; 0.75; 1.0 and 1.25 L ha, in a randomized complete block design with 4 replicates. Productive and qualitative parameters were evaluated. The data were submitted to analysis of variance, using for the comparison of means, the Regression test at 5% of significance. The biostimulant promotes an increase in the accumulation of DM and in the rate of accumulation of forage, leaves and stems of Urochloa hybrid and reduction in the percentage of dead material and the L:S ratio of Urochloa hybrid.


Introduction
Forage production is preponderant in Brazilian livestock as it deals with the most economical and practical way of producing food for most of the Brazilian cattle herd due to the low cost of production provided by pastures when compared to confinement systems (Berchielli et al., 2012).
The use of new fertilizer technologies, such as the use of biostimulants, when applied externally in plants, promote activities similar to phytohormone groups that aid in defense mechanisms, promote growth and development (Neto et al., 2014), act as activators of plant cell metabolism, provide vigor to the immune system, and reactivate physiological processes in the different stages of plant development .
In recent years, some studies have been carried out with the use of biostimulants in annual and perennial plants and presented controversial results in which positive results were found by authors such as Albrecht et al. (2009) in cotton; Castro et al. (2008); Albrecht et al. (2011) and Albrecht et al. (2012) in soybean; Costa et al. (2010) in coffee and De Almeida et al. (2014) in beans. On the other hand, Ferreira et al. (2007) in corn and Rampim et al. (2012) in wheat did not obtain answers with the application of biostimulant.
In Brazil, the expansion of cultivated pasture areas has occurred in large proportions, especially with species of the genus Urochloa, and probably was never equaled by other forages in any other tropical country (Costa et al., 2007). The Urochloa hybrid Convert HD364 is the result of crossing in three generations: Urochloa ruziziensis × Urochloa decumbens cv. Basilik and the crosses of the progenies obtained in the first crossing with Urochloa brizantha cv. Marandu (Argel et al., 2007).
The hybrid grass Convert HD364 combines productivity, resistance and digestibility, since it has wide adaptability to varied climatic and soil conditions (Santos et al., 2015). However, according to Fagundes et al. jas.ccsenet. (2005), ev achieved a The object biostimula

Materia
The experi in the mun 805 m, in t The climat dry winter of annual the vicinit average tem  of the biostimulant was carried out with the aid of a pressurized CO 2 costal spray, equipped with a 2 meter bar and 4 fan-type nozzles, with a volume of 200 L ha -1 being applied.
For the evaluation of the following variables: leaf content and accumulation of macro and micronutrients of the aerial part; forage accumulation (DM-dry mass) and RA (rate of accumulation) of forage; concentration of PB (crude protein), NDF (neutral detergent fiber) and ADF (acid detergent fiber); relation of L:S (leaf:steam) and RA of leaves, stem and dead material and root mass, the forage mass and reference leaf were collected previously to the cuts made in total area, homogenizing and separating the materials at the end of the experiment.
The last fully expanded leaf per plant unit was randomly collected, referred to as the reference leaf, totaling 30 units per plot (Malavolta, 2006). These were placed in paper bags and fed to the forced air circulation oven at a temperature of 65 °C for a period of 72 hours. After drying, the samples were passed through the milling process in Willey mill (2 mm), identified and sent to the Brazilian Laboratory of Agricultural Analysis LTDA (LABRAS), Monte Carmelo, MG.
The methods used for the determination of the macro and micronutrients were: sulfuric digestion of N (Total N), nitro perchloric digestion for Phosphorus (P), Potassium (K), Calcium (Ca), Magnesium (Mg), Sulfur Copper (Cu), Iron (Fe), Manganese (Mn), Zinc (Zn) and incineration to Boro (B) (Malavolta, 2006). For accumulation of macro and micronutrients in the forage, the foliar content of each nutrient was multiplied by the accumulation of leaf DM in kg ha -1 .
For analyzes of forage accumulation and forage RA; the concentrations of CP, NDF and ADF; percentage, accumulation and RA of leaves, stem and dead material and the relation of L:S, the forage mass of Urochloa hybrid was collected in 2 areas delimited within each plot with the aid of a metallic square (1.00 × 0.50 m), which was cut at random, being cut at 15 cm from the ground level. The sample from each plot was homogenized, separated into subsamples and placed in identified plastic bags.
In order to determine the accumulation of forage, the sub sample was first weighed to produce the green mass, then taken to the forced circulation oven at 65 °C for a period of 72 hours, performing the forage weighing (DM). By means of the relationships between green mass and forage, the percentage of forage (% DM) was calculated and from that value the accumulation of forage in kg ha -1 .
After determining the accumulation of forage, the sub-sample was initially milled in a Willey mill (2 mm). Afterwards, they were stored in plastic bags to perform CP, NDF and ADF analysis.
N total determinations were performed according to Kjeldahl semimicro method (Nogueira & Souza, 2005). From the total N values, the CP content was estimated by multiplying it by the conversion factor of 6.25, considering that the proportion of N in the plant proteins is equal to 16% (Campos et al., 2004).
The determination of the lignin content is performed from the ADF concentration, as described by Silva and Queiroz (2006) and the evaluations of the concentrations of NDF followed the protocols suggested by Mertens (2002).
For the evaluation of the morphological composition, the material was fractionated, with the aid of a scissors, in leaves (green slides), stems (stem and leaf sheaths) and dead material. After separation, each fraction was placed in a paper bag, weighed and taken to the forced circulation oven at a temperature of 65 °C for a period of 72 hours. Then they were weighed again. The weighing data were used to calculate the percentage of forage of each morphological component in relation to the total forage of the sub-sample.
From the percentage of each component, the accumulation of forage of leaves, stems and dead material per hectare was calculated.
Accumulation of forage of component = % of Component × Accumulation of forage in kg ha -1 .
For the calculation of relation of L:S, the percentage of leaves was divided by the percentage of stem.
RA was given by the accumulation of forage, total forage accumulation of each morphological component (leaf, stem and dead material) divided by the total number of days of the period.
A collection of roots mass samples was carried out in the water period, in which the entire clump was removed with a pair of scissors, collecting four points with the aid of a root sampling probe, at three depths, forming a sample composed of each depth and conditioning them in identified plastic bags.
The roots were washed, removing all soil and impurities according to Kanno et al. (1999) by storing the root samples in identified paper bags and placing them in a forced air circulation oven at a temperature of 65 °C for a period of 72 hours. After this period the samples were weighed and weighed, estimating root masses (kg m -3 of DM) at depths of 0-10, 10-20, 20-40 and 0-40 cm.
The results were first submitted to the assumptions, homogeneity, heterogeneity and additivity tests, in order to evaluate the residues normality and the homogeneity of the variances, respectively. After that, the data were submitted to analysis of variance. For the evaluation of the effects of biostimulant doses, we used polynomial regressions at 1 and 5% significance.

Results and Discussion
The use of biostimulant promoted interference in the accumulation of DM from forage, leaves and stems of Urochloa hybrid (p > 0.05), where as DM accumulation of dead material was not influenced (p > 0.05) ( Table 2).   The increases in DM accumulations promoted by the use of biostimulants ranged from 12.7 to 40.2% in forage; 15.3 to 38.7% in the leaves and 2.6 to 85.5% in the stems in relation to the control (Table 2), this increase was related to the fact that the biostimulant contains synthetic substances with functions similar to phytohormones, acting in the growth and plant development, since they function as highly specific chemical signals between cells (De Campos et al., 2015). Serciloto et al. (2008) demonstrated an increase in yield of orange pear with the application of foliar biostimulator at 8 mg L -1 when the plants presented 25% of open flowers.
The reduction of the relation of L:S may be related to the greater accumulation of DM in the shoots in relation to the leaves in the treatments in which they used doses of biostimulant ( Figure 3). In addition, the substances present in the biostimulant act to retard the aging of the plant, stimulating the cell division, the development of lateral buds and the caulinar cell stretching. Thus, the increase of the doses of biostimulants contributed to the reduction in the relation of L:S (p < 0.05) (Figure 3). The percentage of leaf, stem and dead material were not influenced (p > 0.05) by the biostimulant (Table 2).
From the data of percentage of leaf, stem and dead material in the time of the waters (Table 2) it was observed that the relation of L:S was similar for all the treatments. For a good development of forage grass species an ideal temperature and precipitation condition is required, the mean values of precipitation and temperature were 200 mm and 25 °C, respectively. In January of 2015, the average temperature was above 25 °C and the average precipitation was below 200 mm, which affected the process of photosynthesis and evapotranspiration, and consequently made the plant processes as absorption and less active transport, thus impairing the accumulation of live material (leaves + stems) (Figure 1).
The critical limit of the relation of L:S is equal to 1.00. In this study, in all treatments, the relation was higher than this limit for the two seasons (Table 2) (Bonfim- Silva et al., 2010). Due to the high precipitation (Figure 1), it offered better conditions to the tillering of the plants, and consequently favored the relation of L:S. The importance of these data is related to forage quality, since the higher relation promotes a better digestibility of the plant material, and as a result, better animal production.     Regarding NDF, all treatments with doses of biostimulants (Table 4) were below the value indicated by Serafim (2010). For this author, in the management of pastures aiming at the production of ruminant animals, obtaining forage with the NDF content above 65% did not promote losses in DM consumption (forage) by cattle, but in a condition below this value gives ruminal microorganisms greater use of nutrients.
In the comparison of the percentage of NDF (Table 4) with those found in the literature, it was found that this was below those recorded by Velásquez et al. (2010) for Urochloa brizantha cv. Marandu, Corrêa et al. (2007) for coastcross grass and Paciullo et al. (2009) for Urochloa decumbens.
The percentage of ADF was found to be between 27 and 28.5%, and it is noted that these values are below that found in the literature, with variations between 30 and 40% for forage pastures (Velásquez et al., 2010;Neres et al., 2011;Silva et al., 2012;Sanches et al., 2015). However, Nussio et al. (2011) concluded that the ADF maximum limit is 30% and in this case all treatments were below this value (Table 4). Since the percentages of ADF represent lignocellulose (lignin and cellulose), that is, the less digestible portion of the cell wall and according to Nussio et al. (2011), the lignin present in the cell walls is what most limits the digestion of fiber, mainly to indicate the maturity of the tissue.
By relating the concentration of ADF and NDF mainly composed of proteins, fats, soluble carbohydrates and pectin, and other water soluble constituents, obtained values (Table 4) afforded the voluntary intake, demonstrating good palatability and digestibility Urochloa hybrid, corroborating with Argel et al. (2007).
The CP grasses and legumes have in their composition a percentage of N non-protein (Santos & Pedroso, 2011), and fertilizers, especially nitrogen, in addition to increasing the production of forage, increase CP content of the fodder, and in some cases, decrease the fiber content, thus contributing to the improvement of Urochloa hybrid quality (Table 4).
The levels of CP found in the present work are above 8% of CP, a minimum considered to meet the requirements of nitrogenous compounds of ruminal microorganisms, and do not compromise the use of the available energy substrates (Lazzarini et al., 2009). Argel et al. (2007) concluded that the adequate CP content for Urochloa hybrid is between 8 and 16%, corroborating with the value observed in the present study (Table 4).
No interference of biostimulant doses (p > 0.05) was observed on the macro and micronutrient contents of Urochloa aerial part (  The N is among the most important factors to determine the level of production per area and its absorption directly influences the CP content of the forage (Lazzarini et al., 2009). In the present study, the N content in Urochloa in the aerial part was in the range of 13 to 20 g kg -1 , and it is noted that no treatment obtained adequate levels of N (Table 5), which was not expected due to high nitrogen fertilization (30 kg ha -1 ) in all treatments. The dose of 1.0 L ha -1 of biostimulant had the leaf content closest to the appropriate one, 12.5 g kg -1 .
In the present work, the two elements mentioned are in the range of 0.8 to 3 g kg -1 and for K of 12 to 30 g kg -1 . content. The foliar contents of the nutrients Ca, Mg and S were within the appropriate range in the aerial part (Table 5), being 3 to 6 g kg -1 ; 1.5 to 4.0 g kg -1 and 0.8 to 2.5 g kg -1 respectively (Perondi et al., 2007).
According to Oliveira et al. (2007) the appropriate values for micronutrients are: B between 10 to 25 mg kg -1 ; Fe between 100 and 487 mg kg -1 ; Mn between 40 to 250 mg kg -1 ; Zn between 20 and 50 mg kg -1 . Analyzing the leaf contents, it was verified that these nutrients were found within the appropriate range (Table 5).
The content of Cu in the forage should be at least 5 mg kg -1 (Sousa et al., 2005), for all dosages desirable nutrient levels were observed (Table 5). Micronutrients are required by plants in small quantities, although the lack of any one can limit plant growth even when all other essential nutrients are present in adequate quantities.
Micronutrients have essential functions that take part in the metabolism of plants, and act mainly as catalysts of various enzymes (Lopes, 1998).
In general, the biostimulant acts directly on root absorption and indirectly, when it has an effect on increasing or reducing the demand for several compounds, including minerals (Vieira & Castro, 2004). In the papers reviewed, it was observed that the substances regulating growth influenced several physiological phenomena related to mineral absorption, such as membrane conductivity and metabolic use of ions (Van Stenveninck, 1976). However, for Albuquerque et al. (2000), plant regulators that inhibit the synthesis of gibberellins and branch growth have influenced the concentration of nutrients in various perennial crops.
The indirect effect of biostimulant on mineral absorption, which occurs through vigor control, was based on the relationship between mineral absorption and growth demand. According to the concepts of Russell (1977), the absorption of nutrients is mainly determined by the metabolic demand of the plant. Phytohormones regulated jas.ccsenet.org Journal of Agricultural Science Vol. 11, No. 6; plant growth by different root management, finding a linear relationship between Ca and K uptake and increment in forage (Richards & Rowe, 1977).
The accumulation of N in the forage of Urochloa hybrid suffered interference (p < 0.05) from the doses of biostimulant and the other nutrients (K, P, Ca, Mg, S, B, Cu, Fe, Mn and Zn) did not suffer biostimulant interference (Table 6). Note. C.V. (%): Coefficient of variation. P-value: ns: not significance to 1% and 5%.
Studies conducted with U. brizantha by Faquin et al. (2000), in Cerrado latosols, showed that the symptoms of P deficiency in forage were the most severe, which did not corroborate with the results found in the present study.
In the analysis performed at the beginning of the experiment it was found that P content in the soil was below the recommended level (Table 1) and when analyzing the leaf contents (Table 5) and the accumulation of P in the plant (Table 6) were within the recommendation range. According to Argel et al. (2007), the use of N by Urochloa hybrid improved the adequate levels of minerals, and may explain the improvement in leaf content of P, since this nutrient was not available to the plant in the form of fertilizer.
The macronutrients were accumulated in the following order, K > N > P > Ca > Mg > S and Fe > Mn > B > Zn > Cu for micronutrients (Table 6), corroborating Braz et al. (2004) for Urochloa.
The biostimulant has the capacity to stimulate root development, increasing the absorption of water and nutrients by the roots, and can also favor the hormonal balance of the plant (Vieira & Castro, 2004). Was made available to the Urochloa hybrid, together with the application of the biostimulant and as it was applied in all plots, even in the control. The progressive increase of the accumulation up to the maximum dose demonstrated that the product potentiated the absorption of N by the plant and made N available to the plant.
No regression models were found that adjusted for the increase in the accumulation of other nutrients (K, P, Ca, Mg, S, B, Cu, Fe, Mn and Zn) in the leaves of Urochloa hybrid, but for all, up to 45.8% over that produced at the control (Table 6). In the results found in the literature, K is the macronutrient of greatest accumulation, followed by N, these nutrients being the most absorbed and accumulated in the vegetal tissue of the plants in the Cerrado region (Pariz et al., 2011;De Mendonça et al., 2014). This corroborated with the accumulation found in the present study (Table 6).

Conclusions
Biostimulants promote an increase in the accumulation of DM and in the rate of accumulation of forage, leaves and stems of Urochloa hybrid Convert HD364 and reduce the percentage of dead material and the relation of L:S of Urochloa hybrid Convert HD364.