Iwona Puzio 
The effect of phytase, cholecalciferol and calcitriol 
on growth of broiler chickens 

The aim of the undertaken research was to determine the influence of separate of joint supplementation of phytase and 1,25(OH)2D3 (calcitriol) to the low-phosphorus and low-calcium diet without vitamin D3 or with its reduced level for the period from 1 to 49 day of life on general growth and development of broiler chickens, growth, development and resistance of osseous system, the level of biologically active substances determining calcium-phosphorus homeostasis and metabolism of bone tissue. The aim of the paper was also to establish whether the supplementation of low-calcium and phosphorus diet with phytase or/and calcitriol in the absence or reduced level of vitamin D3 is a requisite of the same growth and development effects on the organism and particularly on the osseous system as a standard diet in respect of Ca, P and vitamin D3 content.
The research was carried through on broiler chickens in the period from hatching to the 49 day of life. In the experiment I, chickens were divided into 7 groups: control posi-tive receiving diet with vitamin D3, control negative receiving feed without vitamin D3 and 5 experimental groups fed on the diet without vitamin D3 supplemented only with 1,25(OH)2D3 in the amount of 3 µg/kg of feed, with phytase in the amount of 500 or 750 PU/kg of feed or simultaneously 3 µg 1,25(OH)2D3 and 500 or 750 PU of phytase. In the experiment II chickens were divided into 5 groups: positive control receiving feed with proper content of vitamin D3 (starter 2500IU/kg, grower 2000 IU/kg of feed), negative control receiving diet without vitamin D3 and 3 experimental groups fed on the diet with reduced content of vitamin D3 (starter 1250 IU/kg, grower 1000 IU/kg of feed) treated with the sole or simultaneous supplement of 3 µg 1,25(OH)2D3 or 500 PU/kg of phytase. On the 21 day of life of chickens each experimental group was divided into 2 subgroups. One subgroup continued to receive feed with adopted supplement while the other was fed on the diet free of supplements and which did not contain vitamin D3 in the experi-ment I, or with its reduced content in the experiment II. In order to compare the obtained results to parameters received while feeding with diet having standard content of Ca, P and vitamin D3 the group of reference for both experiments was a standard group.
Body weight in 21 and 49 day of life and feed intake in the period 0-21 and 22-49 days were estimated. The ratio feed intake:weight gain was also calculated. On 21 and 49 day of life chickens from each group were slaughtered in the number n = 9-15. The estimation of slaughter analysis was carried through with the determination of warm carcass weight, cooled carcass weight, wings, breast, thigh and shank muscles, giblets and abdominal fat weight. Femora were isolated and their weight, length, mineral den-sity, physical parameters (maximum force, maximum elastic force), geometrical parame-ters (second moment of inertia in horizontal axis, cross-sectional area) and cortical shaft index (cortical thickness, cortical area, cortical index, cortical area index) were meas-ured. 
The percentage content of ash, calcium, phosphorus and magnesium in the bones was also determined. The content of calcium and magnesium was estimated with the use of the method of spectrophotometry of atomic absorption and the content of phosphorus with colometric phosphoric-molibdic method. During the slaughter blood samples were collected in order to estimate the level of hormones and macroelements in plasma. The content of osteocalcin, 1,25(OH)2D3 and 25OHD3 was measured with the aid of radio-immunological methods and the content of calcium, phosphorus ad magnesium by col-orimetric method. The obtained results were submitted to statistical analysis, determin-ing Tukey’s multiple range test. In order to estimate the influence of used supplements on bone mechanical properties the test of 2- and 3-factor variance analysis was per-formed . Statistical results were processed at p ≤ 0,05.
It was concluded that supplementation of phytase and 1,25(OH)2D3 on the diet ex-erted a beneficial influence on performance of chickens both on 21 and 49 day of chicken’s life. During a joint supplementation of 1,25(OH)2D3 and phytase the influence on achieved body weight resulting from the length of time of supplementation was not confirmed. Whereas, the separate addition of one supplement only until 21 day caused a smaller body weight in comparison with a 49-day period of supplementation. A 21-day supplementation of diet with reduced level of vitamin D3 influenced the increase of feed intake and decrease of its intake in older chickens in relation to a 49-day period of treatment. With the absence of vitamin D3 no bigger influence of supplement use duration on feed efficiency was noted either in separate supplementation of 750 PU of phytase or in a joint addition of phytase and calcitriol (irrespective of the level of supplementation). The use phytase and calcitriol provided an adequate bone resistance to the impact of deforming forces. In case of feeding chickens with diet without vitamin D3, phytase and calcitriol had an impact on the increase of bone resistance in the first period of rearing, and in the second period – mainly calcitriol. With reduced content of vitamin D3, increased bone resistance in younger chickens was dependant on the addition of calcitriol while in older birds on the supplementation of the diet with phytase. The addition of phytase greatly influenced the increase of geometrical bone parameters both in the pres-ence and absence of vitamin D3 in the feed. The lack of major influence of addition period duration of the diet without vitamin D3 on values of mechanical parameters on 49 day proves that the use of phytase and calcitriol in the first weeks of life provides a proper organization of bone tissue, resulting from its architecture and degree of its mineralization which makes it resistant to forces effects in the subsequent periods of development. While birds were fed on the diet without vitamin D3 or with its reduced level, the supplementation of 3 µg of calcitriol exerted an important influence on increase of bone mineral density both in 21 and 49 day of chickens’ life. The addition of phytase and calcitriol to the diet provided a proper mineralization of bone nearing the apposition of a mineral fraction obtained as a results of the feeding with adequate content of Ca, P and vitamin D3. Supplementation of calcitriol influenced the reduction of its contents in plasma. The use of phytase and calcitriol did not have a major impact on the contents of osteocalcin on 21 day of chickens’ life. In older birds a 49-day addition of only phytase or calcitriol to the diet with reduced level of vitamin D3 caused the increase of the osteo-calcin level. With the lack of vitamin D3, the increase of level of osteocalcin in plasma took place as a result of 21 day supplementation of phytase and calcitriol. The supplementation of phytase and calcitriol influenced more the content of calcium and phosphorus in plasma while chickens were fed on the diet without vitamin D3. No bigger impact of used supplements on the content of Ca i P in plasma was noted with the reduced level of vitamin D3 in feed. In spite of indicating a positive influence of phytase and calcitriol on mechanical and structural properties of bone, no interaction was confirmed in the effect of both supplements. On the grounds of obtained results it has been concluded that in the condition of deficit or limited amount of calcium, phosphorus and vitamin D3 in the diet, addition of phytase and/or calcitriol makes it possible to obtain beneficial production results and is a requisite of proper development of osseous-skeletal system of broiler chickens in first 49 days of their life.