Effects of dietary oil sources, calcium and phosphorus levels on growth performance, carcass characteristics and bone quality of broiler chickens

ABSTRACT The study investigated the effects of varying dietary calcium (Ca) level and oil sources on the growth performance and carcass quality of broiler chickens. A total of 378, 1-day-old birds (Cobb 500) were fed either 6% palm oil, soybean oil (SO) or linseed oil (LO) in combination with three dietary levels of calcium (1.00%, 1.25% and 1.50%) for 6 weeks. Birds fed SO had higher body weight (BW) compared with those fed LO (p < .05). However, feed efficiency, carcass and bone quality were similar among the oil treatments. Regardless of the oil source, chicken fed diets containing 1.50% of Ca had lower BW compared with those fed 1.00% and 1.25% of Ca. In contrast, birds fed 1.25% of Ca had significantly higher (p < .05) bone quality than those fed 1% of Ca. It can be concluded that increasing the level of calcium up to 1.25% improved bone quality regardless of the type of oil.


Introduction
Dietary oils have high caloric value and thus provide increased energy levels at a lower cost (Lopez-Bote et al. 1997;Baiao & Lara 2005). In addition, oil improves the absorption of oilsoluble vitamins, increases the palatability of rations, reduces pulverulence, increases the efficiency of the consumed energy (Baiao & Lara 2005;Chwen et al. 2013) and also reduces the rate of passage of digesta in the gastrointestinal tract, which gives room for adequate and efficient absorption of the nutrients present in such diet (Baiao & Lara 2005). Furthermore, the fatty acid profile of muscle tissue reflects the dietary fatty acid profile (Abdulla et al. 2015). However, dietary fat could affect mineral metabolism, especially that of magnesium, calcium and zinc (Atteh & Leeson 1983;Leeson & Atteh 1995). This is due to the formation of insoluble soaps between fatty acids and these minerals during digestion, which renders both the fatty acids and these minerals unavailable if the soaps are insoluble (Atteh & Leeson 1983;Leeson & Summers 2005). This can jeopardize the energy value of the oil and hinder the bird's mineral retention and bone and eggshell quality. Thus, addition of fat to poultry diet necessitates the addition of calcium for better poultry performance. The benefits of calcium supplementation on the production performance of poultry have been underscored (Underwood & Suttle 2001;Peters & Mahan 2008). Calcium has important biological functions and must be provided in adequate amounts. Inadequate calcium intake may affect bone mineral content, muscle function and other functions of minerals in the body (Peters & Mahan 2008). Abdulla et al. (2016) found that different dietary levels of calcium differ in their effect on apparent nutrient digestibility. The modern broiler has been intensively selected for higher growth rates and increased feed conversion. Today, broilers are ready for market at 6 weeks of age with a body weight (BW) of 2.6 kg (Santos et al. 2005). Unfortunately, the mineral requirements of broiler chickens, as determined by several organizations 10-20 years ago, may not support optimal chicken performance in today's strain (Ruttanavut & Yamauchi 2010). It is recommended that when high levels of fat are used in poultry diets, calcium and magnesium levels should be increased (Hakansson 1975). Although some researches have been conducted on the effect of dietary oil and National Research Council (NRC) calcium requirement on broiler performance and carcass quality characteristics (Maroufyan et al. 2012;Poorghasemi et al. 2013), information on the effect of dietary oil sources and different levels of calcium is very limited. Thus, the objective of this study was to determine the effect of soybean oil (SO), linseed oil (LO) and palm oil (PO) each at 6% with different levels of calcium and phosphorus on the growth performance, bone quality and carcass characteristics of broiler chicken.

Birds, husbandry and experimental procedure
A total of 378 day-old male broiler chicks (Cobb) were used for this experiment. Upon arrival, the chicks were individually wing banded, weighed and assigned at random to nine dietary treatments. All treatments were replicated six times. Each replicate consisted of seven chicks. After seven days of the rearing period, all birds were vaccinated with infectious bronchitis and Newcastle disease live vaccine against IB and ND through the intraocular route. The infectious bursal disease vaccine was applied on day 14 of the rearing period through the intraocular route. Water and feed were provided ad libitum. The starter and finisher diets were offered to the broiler chickens from 0 to 21 and from 22 to 42 days of age, respectively. The birds were provided different diets with three oil sources, which were LO, SO and PO and three levels of calcium: phosphorus, namely 1:0.5; 1.25:0.63 and 1.5:0.75 (NRC 1994). The diets were formulated to maintain a constant ratio of energy and protein to meet the requirements for the broiler chickens. Feed ingredients and diets were sampled during the preparation of the experimental diets, and the proximate analysis of crude protein, ether extract and ash was determined according to the standard procedure of AOAC (1995). The dietary composition for starter and finisher diets is presented in Tables 1 and 2, respectively.

Growth performance
The individual BW and pen feed intake (FI) were recorded weekly, and live weight gain and feed conversion ratio (FCR) were calculated.

Slaughter procedure and carcass measurements
Upon completion of the feeding experiment, 12 chickens per group were taken as a representative sample and were slaughtered at the Department of Animal Science abattoir, Faculty of Agriculture, Universiti Putra Malaysia. The birds were slaughtered according to a Halal slaughter procedure as outlined in MS 1500: 2009 (Department of Standards Malaysia 2009). The carcasses were dissected manually and the following parameters were recorded: carcass weight and dressing percentage (carcass weight as % of final BW), and weight of breast meat and leg (thigh + shank). Vitamin premix provided the following per kg diet: Vitamin A (retinyl acetate) 10.32 mg, cholecalciferol 0.250 mg, Vitamin E (DL-tocopheryl acetate) 90 mg, Vitamin K 6 mg, cobalamin 0.07 mg, thiamine 7 mg, riboflavin 22 mg, folic acid 3 mg, biotin 0.04 mg, pantothenic acid 35 mg, niacin 120 mg and pyridoxine 12 mg. f Mineral premix provided the following per kg diet: iron 120 mg, manganese 150 mg, copper 15 mg, zinc 120 mg, iodine 1.5 mg, selenium 0.3 mg and cobalt 0.4 mg. g Toxin binder contains natural hydrated sodium calcium aluminium silicates (HSCAS). h The diets were formulated using feed live International software. The ME content of PO, SO and LO was assumed to be 34.63, 35.01 and 35.08 MJ/kg, respectively, based on values determined previously in our laboratory for 21-d-old broilers.

Physical characterization and composition of tibiotarsus
After slaughter, 12 birds from each group were selected and the right tibia of each bird was removed as drumsticks with the flesh intact and frozen. The drumsticks were later thawed, labelled and then immersed in boiling water (100°C) for 10 min. After cooling to room temperature, the drumsticks were defleshed by hand, and the bone cap and the patella were removed. Thereafter, they were air-dried for 24 h at room temperature. The tibiotarsal length was measured with an electronic digital measuring device and the bones were weighed on a precision balance. Prior to breaking, each bone was marked at midpoint, and external diameters were measured. The bone-breaking strength was measured by means of a three-point bending test, using a Universal testing machine (Model 1000R, with 5000N load cell) by recording the amount of force required to break the bone when applied at a constant speed of 10 mm/min and distance between supports of 50 mm. The thickness of the medial and lateral walls was measured at the midpoint mark using a dial caliper. Medullary canal diameter was calculated by subtracting the thicknesses of the medial and lateral walls from the diameter at the diaphysis. The tibiotarsal index was determined using the following formula: (Ziaie et al. 2011) To determine bone ash content, bones were oven-dried at 105°C for 24 h. The samples were weighed after drying (DM), placed in a pre-weighed crucible and ash in a muffle furnace at 600°C for 6 h and then re-weighed (ASH, g/100 g DM). The total phosphorus content of the bone and feed was determined by using an autoanalyser (Lachat Instruments QuikChem 8000 Series FIA + System) and the calcium was measured by atomic absorption spectrometry (Perkin Elmer Analyst 400).

Statistical analysis
The experiment followed a 3 (sources of oil) × 3 (levels of calcium and phosphorus) factorial arrangement in a completely randomized design. All data obtained were analysed using a generalized linear model of SAS (2007). Significant differences  between treatment means were compared using Tukey's test at a probability of less than 0.05.

Growth performance
The BW and FI of birds fed different oil sources, calcium and phosphorous levels are shown in Tables 3 and 4, respectively. No significant difference in FI and FCR was observed among dietary oils. However, supplementation of broiler diets with SO increased BW and weight gain at the sixth week of age compared with LO (p < .05). The lower final BW and weight gain observed in LO birds could be due to lower fat deposition. This observation corroborates the findings of Crespo and Esteve-Garcia (2002) who reported lower abdominal fat deposition in birds fed LO and sunflower oil compared to those fed tallow and olive oils. The authors attributed their observation to higher de novo fatty acid synthesis, which serves as a major means of energy dissipation and contributed to lower fat deposition found in the birds. The higher BW and daily weight gain in SO birds corroborate the findings of Azman et al. (2004) who observed that the final BW and daily weight gain were higher in broilers fed SO compared to that in those fed a mixture of SO and poultry grease. The similarity in FI observed in the current study agrees with the findings of Smith et al. (2003) who observed that supplementation of fish oil, corn oil, animal fat and a blend of animal and vegetables oils did not influence FI but influenced BW gain and FCR in broilers. However, the present results contradict the findings of Fébel et al. (2008) who reported no significant difference in growth performance of the broilers fed diets supplemented with lard, sunflower oil, SO or LO. Regardless of the oil source, there was no significant difference among the calcium: phosphorus levels for FI and FCR, but the BW and weight gain differ at the sixth week of age. Birds fed diets containing 1.5% of Ca had lower BW and weight gain. The decreased BW observed with increasing calcium level could be due to the increase in the formation of insoluble soap between calcium and fatty acids (Shafey 1998;Mutucumarana et al. 2014). Similar findings were reported by Smith et al. (2003) who observed no significant difference in FI and FCR of broilers fed different levels of dietary calcium and phosphorus. The interaction between different sources of oil and calcium levels was not significant (p > .05) for the growth performance of broiler chicken.

Carcass traits
Understanding the factors that affect carcass characteristics and the quality of chicken meat is important in poultry enterprise (Dyubele et al. 2010). The quality and quantity of carcass are determined by genetic and environmental factors. Amongst environmental factors, a significant influence on the quantity of lean is charged to nutrition (Waldenstedt 2006). The effect of dietary oil sources, calcium and phosphorous levels on carcass weight, dressing percentage, leg % and breast % is shown in Table 5. No significant differences were found among oil groups for dressing percentage, leg and breast % as well as carcass weight. The proportions of the economically important carcass portions were not affected by different dietary oil sources. Similar results have been reported by Zollitsch et al. (1997) who observed that supplementation of animal/vegetable oil blend, SO, rapeseed oil and processed oil product did not influence dressing percentage, proportion and the percentage of leg and breast meat. Similarly, Hulan et al. (1988) found no significant difference in dressing percentage, breast, thigh and drumstick muscle contents among turkey fed diet supplemented with LO, rapeseed oil and SO. However, the present results contrast the findings of AlDaraji et al. (2011) who reported significant difference in carcass efficiency of the Japanese quail fed diets supplemented with linseed, fish and sunflower oils. Calcium levels did not affect yield, leg and breast percentage. However, birds fed 1.5% of Ca had lower carcass weight compared with other levels. This result could be due to lower BW observed in birds fed 1.5% of Ca compared with other levels. This observation is in line with the report of Johnson and Karunajeewa (1985) who observed no significant difference in the dressing percentage of broilers fed different levels of calcium. Contrary to the present observation, Talpur et al. (2012) reported that dressing percentage was higher in broiler chicken fed 10 g/kg of calcium compared with those fed 20 and 30 g/kg DM. The interaction between different sources of oil and calcium levels was not significant (p > .05) for carcass quality of broiler chicken.

Bone characteristics
Leg weakness, lameness and other bone abnormalities connected with various metabolic disorders are continuing problems in rapidly growing carcass quality of chickens, leading to considerable production losses and having negative effects on bird welfare (Venäläinen et al. 2006;Waldenstedt 2006). The effect of dietary oil sources, and calcium and phosphorous levels on physical characterization, bone breaking strength and ash of tibiotarsus is presented in Table 6. Chicks fed diets supplemented with LO had higher bone breaking strength (N), ash percentage and weight of tibia compared to those fed diets supplemented with PO and SO, but the differences were not statistically significant. In line with the current findings, Atteh and Leeson (1984) reported significant (p < .05) reduction Means within the same row for each parameter with different superscripts are significantly different (p < .05). in bone ash and bone calcium content of chicks fed diets supplemented with palmitic acid compared with oleic acid. The authors attributed this to the fact that calcium soaps of dietary unsaturated free fatty acids (oleic acid, C18:1) were absorbed by the bird as opposed to calcium soaps formed with saturated free fatty acids (palmitic acid, C16:0) in postpeak laying hens and broilers. Similarly, Lau et al. (2010), Lau et al. (2013) and Watkins et al. (2000) observed consistent and reproducible beneficial effects of omega-3 fatty acids on bone metabolism and bone/joint diseases. Furthermore, Corwin (2003) found that high-oil diets, especially those rich in saturated fatty acids, may contribute to reduced bone density and increased fracture risk, in older as well as younger people. Regardless of the oil sources, there was significant (p < .05) increase in thickness of the lateral wall, tibiotarsal index, bone breaking strength and bone ash of chicks fed diets containing 1.25% of Ca compared with 1.00% of Ca. Earlier studies observed increased tibia ash or mineral contents with increasing dietary Ca and available phosphorus up to the highest available phosphorus content (5.0 g/kg) (Nelson et al. 1990;Huyghebaert 1996;Onyango et al. 2003). This observation is in agreement with the findings of Venäläinen et al. (2006) which showed that increasing dietary Ca and available P in diet of broiler chicken significantly increased tibia ash content but did not affect tibia breaking strength. Coto et al. (2008) found that bone development was improved by increasing phosphorus and calcium levels. There was no significant interaction (p > .05) between the source of oil and calcium levels on all bone characteristics except thickness of the medial wall of tibia.

Conclusion
The results of this study demonstrated that the addition of SO to broiler diets was more effective in increasing BW at the sixth week compared to LO. However, FI and FCR did not differ among the three oil sources. Irrespective of the oil source, 1.5% of Ca had a detrimental effect on BW gain. No difference in FI and feed efficiency was observed among the calcium levels. Oil source and calcium levels did not influence carcass characteristics. Dietary oil source did not affect bone characteristics except Ca and P composition. The 1.25% of Ca had highest tibiotarsal index, bone breaking strength, thickness of the lateral wall, tibia ash, and Ca and P content, regardless of the oil source.

Disclosure statement
No potential conflict of interest was reported by the authors.

Funding
This project was supported by Long-Term Research Grant Scheme (LRGS) -Food Security entitled 'Enhancing the competitiveness and sustainability of the poultry industry for food security' from the Ministry of Higher Education, Malaysia. Means within the same row for each parameter with different superscripts are significantly different (p < .05).