Aim: The aim of this study was to investigate the effects of phytase which was laboratory produced by Aspergillus foetidus on the growth performance, mineral retention, and bone traits of broilers fed with low dietary calcium and phosphorus.

Materials and Methods: The extracellular phytase enzyme secreted into the crude filtrate was concentrated by ammonium sulfate precipitation to obtain an activity of 500 phytase units (FTU). A total of 90 1-day-old chicks (Cobb 500) were randomly divided into three treatment groups with five replicates having six birds each. Dietary treatment, T1, was with 0.45% non-phytate P (NPP) during starter and 0.40% during finisher phase with 1% Ca. Dietary treatment, T2, had 0.37% NPP during starter and 0.32% in finisher phase with 1% Ca and supplemental lab phytase at 500 FTU/kg. Dietary treatment, T3, was similar to T2 with a lower Ca of 0.8%.

Results: There was no significant difference among the dietary treatments with regard to body weight gain, feed intake, feed conversion ratio, and Ca retention (p>0.05). However, a significant improvement in retention of P by birds was observed in phytase supplemental groups T2 and T3 (p<0.05). Dry weight of tibia (2.58-2.78 g/kg live weight) and ash content (39.7- 41.8%) was comparable among treatments. A similar trend was observed for bone Ca, P, and Mn content.

Conclusion: The study indicated that 500 FTU/kg phytase can be effectively supplemented in a broiler diet with low phosphorus (0.37% in starter and 0.32% NPP in finisher diet) and low calcium (0.8% in diet) for better growth performance and with successful replacement of dietary P by 0.08 % and reduced P excretion into the environment in broiler chicken.

Keywords: broiler, calcium, phosphorus, phytase.

1. Dersjant, L.Y., Awati, A., Schulze, H. and Partridge, G. (2014) Phytase non ruminant animal nutrition: A critical review on phytase activities in the gastrointestinal tract and influencing factors. J. Sci. Food Agric., 95: 878-896. [Crossref] [PubMed] [PMC]

2. Ravindran, V., Cabahug, S., Ravindran, G. and Bryden, W.L. (1999) Influence of microbial phytase on apparent ileal amino acid digestibility in feedstuffs for broilers. Poult. Sci., 78: 699-706. [Crossref] [PubMed]

3. Murai, A., Kobayashi, T., Okada, T. and Okamura, J. (2002) Improvement of growth and nutritive value in chicks with non-genetically modified phytase product from Aspergillus niger. Br. Poult. Sci., 43: 687-695. [Crossref] [PubMed]

4. Cowieson, A.J. and Adeola, O. (2005) Carbohydrases, proteases and phytase have an additive beneficial effect in nutritionally marginal diets for broiler chickens. Poult. Sci., 84: 1860-1867. [Crossref] [PubMed]

5. Butani, J.B. and Parnerkar, S. (2015) Role of microbial phytase in broiler nutrition-a review. J. Livest. Sci., 6: 113-118.

6. Rashti, M.G., Shariatmadari, F., Torshizi, M.A.K. and Asli, M.M. (2016) Effects of dietary propionic acid, sodium citrate, and phytase on growth performance, mineral digestibility, and tibia properties in broilers. J. Appl. Anim. Res., 44(1): 370-375. [Crossref]

7. Ravindran, V., Morel, P.C.H., Partridge, G.G., Hruby, M. and Sands, J.S. (2006) Influence of an E. coli-derived phytase on nutrient utilization in broilers fed starter diets containing varying concentrations of phytic acid. Poult. Sci., 85:82-89. [Crossref] [PubMed]

8. Selle, P.H., Ravindran, V. and Partridge, G.G. (2009) Beneficial effects of xylanase and/or phytase inclusions on ileal amino acid digestibility, energy utilisation, mineral retention and growth performance in wheat-based broiler diets. Anim. Feed Sci. Technol., 53: 303-313. [Crossref]

9. Simons, P.C.M., Versteegh, H.A.J., Jongbloed, A.W., Kemme, P.A., Slump, P., Bos, K.D., Wolters, M.G.E., Beudeker, R.F. and Verschooor, G.J. (1990) Improvement of phosphorus availability by microbial phytase in broilers and pigs. Br. J. Nutr., 64: 525-540. [Crossref] [PubMed]

10. Schlegel, P., Sauvant, D. and Jondreville, C. (2013) Bioavailability of Zn sources and their interaction with phytases in broilers and piglets. Animal, 7(1): 47-59. [Crossref] [PubMed]

11. NRC. (1994) Nutrient Requirements of Poultry. 9th Rev. ed. The National Academies Press, Washington, DC.

12. Angel, R., Tamim, N.M., Applegate, T.J., Dhandu, A.S. and Ellestad, L.E. (2002) Phytic acid chemistry: In?uence on phytin-phosphorus availability and phytase ef?cacy. J Appl. Poult. Res., 11: 471-480. [Crossref]

13. Tamim, N.M. and Angel, R. (2003) Phytate phosphorus hydrolysis as influenced by dietary calcium and micro-mineral source in broiler diets. J. Agric. Food Chem., 51: 4687-4693. [Crossref] [PubMed]

14. Amerah, A.M., Plumstead, P.W., Barnard, L.P. and Kumar, A. (2014) Effect of calcium level and phytase addition on ileal phytate degradation and amino acid digestibility of broilers fed corn-based diets. Poult. Sci., 93: 906-915. [Crossref]

15. Zaefarian, F., Romero, J.F. and Ravindran, V. (2013) Influence of a microbial phytase on the performance and the utilization of energy, crude protein and fatty acids of young broilers fed on phosphorus-adequate maize-and wheat-based diets. Br. Poult. Sci., 54: 653-660. [Crossref] [PubMed]

16. Powell, S., Bidner, T.D. and Southern, L.L. (2011) Phytase supplementation improved growth performance and bone characteristics in broilers fed varying levels of dietary calcium. Poult. Sci., 90: 604-608. [Crossref] [PubMed]

17. Delezie, E., Bierman, K., Nollet, L. and Maertens, I. (2015) Impacts of calcium and phosphorus concentration, their ratio and phytase supplementation level on growth performance, footpad lesions and hock burn of broiler chickens. J. Appl. Poult. Res., 24: 115-126. [Crossref]

18. Rama Rao, S.V., Raju, M.V.L.N., Reddy, M.R. and Pavani, P. (2006) Interaction between dietary calcium and non-phytate phosphorus levels on growth, bone mineralization and mineral excretion in commercial broilers. Anim. Feed Sci. Technol., 131: 135-150. [Crossref]

19. Walk, C.L., Bedford, M.R. and Mcelroy, A.P. (2012) Influence of limestone and phytase on broiler performance, gastrointestinal pH, and apparent ileal nutrient digestibility. Poult. Sci., 91: 1371-1378. [Crossref] [PubMed]

20. Lalpanmawia, H., Elangovan, A.V., Sridhar, M., Shet, D., Ajith, S. and Pal, D.T. (2014) Efficacy of phytase on growth performance, nutrient utilization and bone mineralization in broiler chicken. Anim. Feed Sci. Technol., 192: 81-89. [Crossref]

21. Haugh, W. and Lantzsch, H.J. (1983) Sensitive method for the rapid determination of phytate in cereals and cereal products. J. Sci. Food Agric., 34: 1423-26. [Crossref]

22. Kim, T.W. and Lei, X.G. (2005) An improved method for a rapid determination of phytase activity in animal feed. J. Anim. Sci., 83: 1062-1067. [Crossref] [PubMed]

23. Selle, P.H. and Ravindran, V. (2007) Microbial phytase in poultry nutrition. Anim. Feed Sci. Technol., 135: 1-41. [Crossref]

24. Ahmad, T., Rasool, S., Sarwar, M., Haq, A.V. and Hasan, Z.U. (2000) Effect of microbial phytase produced from a fungus Aspergillus niger on bioavailability of phosphorus and calcium in broiler chickens. Anim. Feed Sci. Technol., 83: 103-114. [Crossref]

25. Lan, G., Abdullah, N., Jalaludin, S. and Hoa, Y.W. (2012) Effects of freeze-dried Mitsuokella jalaludinii culture and natuphos phytase supplementation on the performance and nutrient utilization of broiler chickens. J. Sci. Food Agric., 92: 266-273. [Crossref] [PubMed]

26. Manobhavan, M., Elangovan, A.V., Sridhar, M., Shet, D., Ajith, S., Pal, D.T. and Gowda, N.K.S. (2015) Effect of super dosing of phytase on growth performance, ileal digestibility and bone characteristics in broilers fed corn-soya-based diets. J. Anim. Physiol. Anim. Nutr., 100(1): 93-100. [Crossref] [PubMed]

27. Sebastian, S., Touchburn, S.P., Chavez, E.R. and Lague, P.C. (1996) The effects of supplemental microbial phytase on the performance and utilization of dietary calcium, phosphorus, copper, and zinc in broiler chickens fed corn-soybean diets. Poult. Sci., 75: 729-36. [Crossref]

28. Beiki, M., Hashemi, S.M. and Yaghoobfar, A. (2013) The use of phytase and low phosphorus levels in broiler diets with different metabolizable energy levels. J. Anim. Poult. Sci., 2(2): 48-54.

29. Akter, M., Graham, H. and Iji, P.A. (2016) Response of broiler chickens to different levels of calcium, non phytate phosphorus and phytase. Br. Poult. Sci., 57: 799-809. [Crossref] [PubMed]

30. Rutherfurd, S.M., Chung, T.K., Thomas, D.V., Zou, M.L. and Moughan, P.J. (2012) Effect of a novel phytase on growth performance, apparent metabolizable energy and the availability of minerals and amino acid in a low phosphorus corn-soybean meal diet for broilers. Poult. Sci., 91: 1118-1127. [Crossref]

31. Liu, B., Rafiq, A., Tzeng, Y. and Rob, A. (1998) The induction and characterization of phytase and beyond. Enzym. Microb. Tech., 22: 415-424. [Crossref]

32. Chung, T.K., Rutherfurd, S.M., Thomas, D.V. and Moughan, P.J. (2013) Effect of two microbial phytases on mineral availability and retention and bone mineral density in low-phosphorus diets for broilers. Br. Poult. Sci., 54: 362-373. [Crossref]

33. Singh, A., Walk, C.L., Ghosh, T.K., Bedford, M.R. and Haldar, S. (2013) Effect of a novel microbial phytase on production performance and tibia mineral concentration in broiler chickens given low-calcium diets. Br. Poult. Sci., 54: 206-215. [Crossref]

34. Kiarie, E., Woyengo, T. and Nyachoti, C.M. (2015) Efficacy of new 6-phytase from Buttiauxella spp. on growth performance and nutrient retention in broiler chickens fed corn-soybean meal-based diets. Asian Australas. J. Anim. Sci., 28: 1479-1487. [Crossref] [PubMed] [PMC]

35. Qian, H., Kornegay, E.T. and Denbow, D.M. (1996) Phosphorus equivalence of microbial phytase in turkey diets as influenced by calcium to phosphorus levels. Poult. Sci., 75: 69-81. [Crossref] [PubMed]

36. Rama Rao, S.V., Reddy, V.R. and Reddy, R.S. (1999) Enhancement of phytate phosphorus availability in the diets of commercial broilers and layers. Anim. Feed Sci. Technol., 79: 211-222. [Crossref]

37. Qian, H., Kornegay, E.T. and Denbow, D.M. (1997) Utilization of phytate phosphorus and calcium as influenced by microbial phytase, cholecalciferol, and the calcium: Total phosphorus ratio in broiler diets. Poult. Sci., 76: 37-46. [Crossref]

38. Applegate, T.J., Angel, R. and Classen, H.L. (2003) Effect of dietary calcium, 25-hydroxycholecalciferol, or bird strain on small intestinal phytase activity in broiler chickens. Poult. Sci., 82: 1140-1148. [Crossref] [PubMed]

39. Kim, J.H., Han, G.P., Shin, J.E. and Kil, D.Y. (2017) Effect of dietary calcium concentration in phytase containing diets on growth performance, bone mineralization, litter quality and footpad dermatitis score in broiler chickens. Anim. Feed Sci. Technol., 229: 13-18. [Crossref]

40. Tamim, N.M., Angel, R. and Christman, M. (2004) Influence of dietary calcium and phytase on phytate phosphorus hydrolysis in broiler chicken. Poult. Sci., 83: 1358-1367. [Crossref] [PubMed]

41. Wilkinson, S.J., Selle, P.H., Bedford, M.R. and Cowienson, A.J. (2014) Separate feeding of calcium improves performance and nutrient digestibility in broiler chicks. Anim. Prod. Sci., 54: 172-178. [Crossref]

42. Li, W., Angel, R., Kim, S.W., Brady, K., Yu, S. and Plumstead, P.W. (2016) Impacts of dietary calcium, phytate, and nonphytate phosphorus concentrations in the presence or absence of phytase on inositol hexakisphosphate (IP6) degradation in different segments of broilers digestive tract. Poult. Sci., 95: 581-589. [Crossref]

43. Zeller, E., Schollenberger, M., Witzig, M., Shastak, Y., Kuhn, I., Hoelzle, E. and Rodehutscord, M. (2015) Interaction between supplemented mineral phosphorous and phytase on phytate hydrolysis and inositol phosphates in the small intestine of broilers. Poult. Sci., 94: 1018-1029. [Crossref] [PubMed]

44. Yan, F., Kersey, J.H., Fritts, C.A. and Waldroup, P.W. (2006) Effects of phytase supplementation on the calcium requirement of broiler chickens. Int. J. Poult. Sci., 5: 112-120. [Crossref]

---