INTRODUCTION:

The use of metabolites of vitamin D on broiler diets prevents skeletal problems and promotes bone quality to support gain of muscular mass in broiler strains of rapid growth, especially on birds with bone quality problems, as tibial dyschondroplasia, that affects the performance (^{BRITO et al., 2010}; ^{GARCIA et al., 2013}). The more available metabolites to use as additives, are hydroxycholecalciferol (25(OH)D₃) and 1,25-dihydroxyvitamin D₃ or calcitriol (1,25(OH)₂D₃) (^{APPLEGATE & ANGEL, 2005}).

In all animals, the ingested vitamin D₃ is metabolized in 25(OH)D₃ on the liver, and then, it is metabolized in 1,25(OH)₂D₃ (the active metabolite) on the kidneys (^{WHITEHEAD, 2002}). Besides 1,25(OH)₂D₃ could be obtained from botanical sources, as dry leaves of Solanum glaucophyllum (^{BOLAND et al., 2003}), also known as Solanum malacoxylon or Solanum glaucum, a common bush from temperate zone of South America. However, the aqueous extract could be considered toxic due to high levels of calcium, magnesium and phosphorus ions, which leads to depressed of bone resorption. Therefore, desirable bioactive form could be obtained from the salt-free extract of Solanum glaucophyllum (^{LLOYD et al., 1975}).

The supplement industry works with a safety margin of two to ten times greater than the actual need of the birds, to guarantee the minimum supplementation of the nutrients after the manufacturing physical processes, transportation and diets storage (^{COELHO et al, 1995}; ^{FÉLIX et al. 2009)}. However, ^{BRITO et al. (2010}) did not recognize the advantage of using high levels of vitamin D₃ on performance of broilers, as well as ^{ALAHYARI-SHAHRASB et al. (2012}) suggested that it has been possible to reduce the amount of some vitamins in the premix for finisher’s diets by 33% without affecting the performance and metabolic characteristics.

The bone quality is assessed by measuring bone’s density, resistance to breakage and percentage of minerals in the bones, through mechanical, physical and chemical features. If the features of bone quality go along with the production rates in the flocks, in satisfactory way, then, the supplementation with metabolites could be positively evaluated.

Studies related with the use of vitamin D₃ metabolites can help companies with integration systems, which would handle mineral and vitamin supplements, to reduce the amount of vitamin D₃ by association with active metabolites; and therefore, enable cost reduction without loss on performance, carcass yield and bone quality of the flocks of chickens. Based on these considerations, this study aimed to evaluate the effect of the vitamin D₃ reduction in the premix and, the 1,25(OH)₂D₃-glycoside supplementation on performance, bone quality and carcass yield from 7 to 42d broilers.

MATERIALS AND METHODS:

RESULTS:

DISCUSSION:

When 1,25(OH)₂D₃-glycoside was used as the single source of vitamin D₃, many performance, yield and bone quality variables of broilers Cobb500^® at 42 days was extremely impaired. Similar results were previously reported, low values of weight gain associated to moderate hypercalcemia were reported in six weeks broilers supplemented with 1,25(OH)₂D₃ (^{RENNIE et al., 1995}; ^{RENNIE et al., 1996}). In order to justify the occurrence of the poor results, it would be necessary to perform analysis such as the blood and organs metabolite profiles, 1,25(OH)₂D_3, and others related to the bone formation process, for the purpose to know if the use of fixed quantity (50g of the product per ton of feed), would provide lower levels of vitamin D₃ than required within each stage of development.

The use of 100, 75, 50 or 25% of vitamin D_3, in addition of 1,25(OH)₂D₃-glycoside, did not affected the performance, and it was close to the Cobb500^® standards. In a similar way, vitamins reduction in 33% of the finishing diet premix for broilers rising in floor, did not affect the performance at slaughter age (^{ALAHYARI-SHAHRASB et al., 2012}). Additionally, ^{COLET et al. (2015}) studied vitamin reductions in broiler diets and, reported that there was no need for further vitamin supplementation when mineral levels had been balanced. ^{VIEITES et al. (2014}) used 1,25 (OH)₂D_3-glycoside metabolite to supplement broiler in a quantity five times higher (250 g of Panbonis® ton^-1) than that used in the present study and, did not observe significant differences in the performance of 42-d old broiler chickens. According to ^{FÉLIX et al. (2009)}, the vitamin levels used by the industry are 10 to 25 times higher than the recommended by the ^{NRC (1994)}, and once the premix used was formulated based on a commercial brand, the vitamin levels were enough to guarantee the standard performance of the broiler strain used on this study. Calcidiol supplementation plus different levels of vitamin D₃ reduces the mortality rate (^{MICHALCZUK et al., 2010}). Still, in the present research, calcitriol supplementation did not affect the viability rate.

The carcass and parts yield (wing, breast and thigh + drumstick) did not differ from control. Similar results were reported by ^{VIEITES et al. (2012}). The mechanisms of the 1,25 (OH)₂D₃ on muscles of chickens are still to be determined, but is known that vitamin D is necessary for skeletal muscle development and integrity (^{DOMINGUES-FARIA et al., 2017}). In mice, ^{GIRGIS et al. (2015}) observed that vitamin D deficiency reduced muscle fiber, found direct effect of vitamin D on the muscle function and corroborated the presence of vitamin D receptors in this tissue. Reduction or withdrawal of some vitamins and minerals in finishing diets did not affect the composition of carcasses of broiler chickens; however, withdrawal of vitamins in the premix during growth period leads to lower carcass yield (^{MAIORKA et al., 2002}; ^{KHAJALI et al., 2006}).

The opposite was observed by KORVER (2005) cited by ^{VIEITES et al. (2014}), he reported a better carcass and breast yield in 42-d broilers by vitamin D₃ plus 25(OH)D₃ supplemented diets. Also, ^{SAUNDERS-BLADES & KORVER (2006}) observed advantages 25(OH)D₃ metabolite on breast muscle development, yield and meat quality in 42-day-old broilers, similar data were reported by ^{BRITO et al. (2010}) because they supplemented with high levels of 25(OH)D₃, and observed higher carcass yield compared to the effect of vitamin D₃.

Bone quality of the tibia was affected and tibia length, the weight were lower when the birds did not receive vitamin D₃ in the diet and the single source of vitamin D was the metabolite 1,25(OH)₂D­₃ glycoside. Additionally; although, weight/length ratio had not been affected the breaking strength of the bones was lower in those birds, which could be related to the smaller mineral matter of their bones. The stiffness of the bone tissue results from the deposition of calcium and phosphorus, in the form of hydroxyapatite, during the process of bone mineralization. These two minerals make up about 70% of the bone composition; the remaining 30% are composed of organic matter, mainly collagen. Active vitamin D₃ acts on the bone, stimulating protein synthesis by osteoblasts and participating in matrix mineralization (^{BORGES et al., 2010}; ^{SOUZA, 2012}). Bones undergo several adaptations throughout development, and mineral content is the main determinant of the bones mechanical properties, in several species, as that, the higher the mineral content the greater the bone strength (^{CURREY, 2003}). This observation explains the effect occurred in the treatment with the use of metabolite as a single source of vitamin D, because the lower percentage of ash was accompanied by the lower mean breaking strength. Conversely, ^{TATARA et al. (2011}) studied the effect of 25(OH)D₃ supplementation and did not observe significant differences in turkey tibia length up to 20 weeks of age.

Supplementation of 1,25(OH)₂D₃-glycoside as a single source of vitamin D promoted a reduction of the concentration of minerals in the broiler’s tibia (from 47.86% to 40.14%). However, it was still within the acceptable percentages of broiler’s bone ash (40 - 45%) in birds receiving vitamin D₃ supplementation (^{AOAC, 1995}). Similar results were reported by ^{GARCIA et al. (2013}) when they used 1,25 (OH)₂D₃ in broiler diets, and obtained a percentage of ash around 44%.

When 1,25(OH)₂D₃-glycoside was used as the single source of vitamin D, the percentage of calcium deposited in the bone did not differ from the control treatment, reinforcing the role of 1,25(OH)₂D₃ in the physiological regulation of intestinal absorption of Ca, as it was supported by reports of ^{SILVERTHORN (2010}), that observed that 1,25(OH)₂D₃ could increase active transport of Ca by up to 30%. According to ^{NORMAN & HENRY (2007}), when 1,25(OH)₂D₃ was given to vitamin D-deficient chicks it was observed that calcium transport reached peak rates between 12 and 14h, maintaining adequate circulating levels and decreasing mobilization of calcium from bones. The 1,25(OH)₂D₃ plays a significant role in the regulation of active calcium transport, ensuring maintenance of plasma levels and reducing the mobilization of this mineral in bones when the nutrition and vitamin and mineral supplementation are correct (^{SILVERTHORN, 2010}).

^{TATARA et al. (2011}) studied the effect of 25(OH)D₃ supplementation, nor did they observe differences between tibial mineral volume density (measured by computed tomography and the Somatom Emotion Siemens apparatus) in turkeys up to 20 weeks of age. However, in the study by ^{GARCIA et al. (2013}), supplementation with Panbonis^® at a concentration of 0.250g/kg diet led to higher weight/length ratio values and greater breaking strength (approximately 336.46N) when compared to the present results.

In the present study, no histological evaluation of the bones was performed to the effect of supplementation with the metabolite on the bone structure. However, the use of decreasing concentrations (100, 75, 50 or 25%) of vitamin D₃, that is, the reduction of up to 75% of the inclusion of vitamin D3 in the premix plus the 1,25(OH)₂D₃ glycoside supplementation, did not alter any of bone quality variables of the broilers that were evaluated in the present study.

For the purposes of the present research it was concluded that the performance, carcass weight and bone quality of broiler chickens are affected by the non-inclusion of Vitamin D₃ in the premix; although, supplemented with 50g/ton of 1,25 (OH)₂D₃ glycoside. A reduction of vitamin D₃ from the premix up to 75% when the inclusion of 1,25(OH)₂D₃-glycoside maintains the performance and bone quality of the male broilers of the Cobb500^® lineage. Carcass yield and noble cuts are not influenced by the reduction of vitamin D₃ level of premix and supplementation with 1,25(OH)₂D₃ glycoside. However, the use of 1,25(OH)₂D₃ glycoside as the single source of D in the amount tested is not recommended for male broilers Cobb500^®.