Abstract: The purpose of the present study was to examine the effect of dietary Total Phosphorus (TP) on performance, serum P, bone morphometric measurements, phosphorus (P) and calcium (Ca) retention and P excretion of broilers from 3 to 7 weeks of age. The grower diets (3-6 weeks) contained 5.0, 5.5, 6.0 and 6.5 g kg-1 TP (2.4, 2.9, 3.4 and 3.9 g kg-1 calculated non-phytate P (npP), respectively); each diet was fed to 6 replicate pens. During finishing period (6-7 weeks), three pens received the same TP level as in the growing period, while the other 3 pens were fed a diet without any inorganic P supplementation (3.8 TP, 1.3 npP g kg-1). Decreasing dietary TP from 6.5 to 5.0 g kg-1 during the growing period or removing dicalcium phosphate completely from the finisher diets had no significant effect on feed intake, body weight gain, breast muscle yield and Feed Conversion Ratio (FCR). Serum P was affected by the amount of P in the diet. Tibiae and femora ash percentages were differed significantly between diets and sex. Phosphorus intake and excretion were functions of the amount of the P in the diet, reducing or omitting P in the diets markedly reduced fecal P. In conclusion, TP levels needed by broilers to maintain growth and feed utilization were less than those needed for maximum tibia ash; TP could be reduced to 5.0 (2.4 npP) g kg-1 from during the growing period and omitted from the withdrawal diets without adverse effects on live performance or skeletal integrity.
INTRODUCTION
About 50-80% of the P in grain cereal and oilseed is bound to phytic acid (hexaphosphate of myo-inositol); the organically bound form of P is called Phytate Phosphorus (PP) (Eeckhout and DePaepe, 1994). Despite the amount of literature published on the role of PP in poultry nutrition, controversy still exists about the ability of poultry to utilize PP. The availability of PP usually ranges from 0 to 50% (Nelson, 1976; Edwards et al., 1989; Mohammed et al., 1991). Nutritionists routinely include a margin of safety for phosphorus when they formulate poultry diets. The portion of P that exceeds the requirement will eventually pass through the digestive tract and be excreted in feces creating concerns regarding environmental contamination. This is particularly true in areas of intensive poultry production; therefore, excess P is to be minimized (Cowieson et al., 2004; Musapuor et al., 2006; Manangi and Coon, 2008; Liem et al., 2009; Bingol et al., 2009).
It should be noted that many investigators have utilized maximum bone ash as the criteria for determining the chick’s phosphorus requirements. The question remains regarding whether or not maximum calcification is necessary for a good carcass in the poultry industry. It would seem reasonable to seek a phosphorus level which yields maximum growth, maximum feed efficiency and adequate but not necessarily maximum, bone ash (Edwards, 1983; Ravindran et al., 1995). Several reports have shown that 5 g kg-1 TP is required for maximum growth. However, a higher level was required for maximum bone ash (Dhandu and Angel, 2003; Abas et al., 2011).
The evidence suggests that P excretion in manure could be decreased by formulating grower and finisher diets with reduced concentration of inorganic P. The purpose of the present study was to (1) examine the effect of different levels of dietary TP on performance, P excretion; P and Ca retention (2) examine the effects of TP and sex on femora and tibiae ash and femoral morphologic integrity of broilers.
MATERIALS AND METHODS
Animals and treatments: Day-old, mixed chicks (Ross x Arbor Acres) were placed into 24 pens (2.6 m2 pen-1) in an open-sided poultry house with partial environment control. At 3 week, birds were weighed on a pen basis and stocking density adjusted to 22 birds per pen. In the course of the experiment a metabolism trial was performed with 48 birds housed 2 per cage. The birds were exposed to 23 h light during the trial period.
Chicks were fed a standard corn-soybean meal starter mash ad libitum to 3 weeks of age. Ingredients were analyzed for TP and Ca content (AOAC, 1984) and the adjusted values were used to formulate the experimental diets. The grower diets contained 5.0, 5.5, 6.0 and 6.5 g kg-1 TP (2.4, 2.9, 3.4 and 3.9 g kg-1 calculated npP, respectively) as presented in Table 1. Each diet was fed to 6 replicate pens (3-6 weeks). From 6 to 7 weeks each treatment was divided into two groups, three pens received the same TP level as in the growing period, treatments 1 to 4 (5.0, 5.5, 6.0 and 6.5 g kg-1 TP) while the other 3 pens were fed a diet without inorganic phosphorus supplementation (treatments 5 to 8) and contained 4.0, 1.3 g kg-1 TP, np P, respectively (Table 1). Metabolism trial conducted in conjunction with experiment 1, the diets were fed to birds maintained in wire-floored battery pens, with 2 birds per pen. Each diet was fed to 6 replicate pens (3-6 weeks) and 3 pens (6-7 weeks).
Measurements: Body weight and feed consumption were recorded weekly by pen and FCR computed at 6 and 7 weeks of age. At both ages one female and one male was sampled from each pen. Blood samples were collected by cardiac puncture for serum phosphorus (Baginski et al., 1967) and after euthanasia, the breast muscle was dissected and weighed (Uittenboogaart and Gerrits, 1982). The left limb was identified and the middle toe (between second and third tarsal) was removed from each sample. Toes were cleaned, dried overnight at 100°C and ash determined (AOAC, 2000). The left limbs were cleaned, tibiae and femora were obtained, dried at 100°C and fat was extracted with hexane. Tibiae were ashed and femora were used to determine the morphometric measurements then were ashed (Crespo et al., 2000). The metabolism trial determined TP excretion (3-6 weeks) and Ca and P retention at 6 weeks. Birds (6 replications per each diet) were housed (2 birds/cage) in 24 cages with wire bottoms. Birds had free access to feed and water throughout the trial period. All diets were supplemented with 3 g kg-1 chromic oxide as an analytical marker and birds were fed chromium oxide mashed feed for 5 days and excreta was collected to determine Ca and P retention (Edwards and Gillis, 1959). Feed and excreta samples were dried at 60°C until constant weight and were ground, Ca and P concentrations in feed and excreta were determined by inductive coupled plasma (ACP) analysis (Smith, 1965) and Cr concentration was determined (Bolin et al., 1952). Percent Ca and P retention was calculated (Scott et al., 1976).
| Table 1: | Dietary ingredients (g kg-1) and chemical composition of the broiler grower (3-6 weeks) and finisher (6-7 weeks) diets |
| 1Vitamin mix provided per kg diet using roche broiler premix: Retinyl acetate, 3.41 mg; cholecalciferol, 0.07 mg; DL-α-tocopheryl acetate, 27.5 mg; menadione sodium bisulphate, 6 mg; riboflavin, 7.7 mg; niacin, 44 mg; pantothenic acid, cyanocobalamin, 0.02; choline 496 mg; folic acid, 1.32 mg; pyridoxine HCl, 4.82 mg; thiamine mononitrate, 2.16 mg; D-biotin, 0.11 mg, 2Mineral mix provided per kg diet using Mar-Jac Poultry Trace Mineral Mix: manganese, 67 mg; zinc, 54 mg; copper, 2 mg; iodine, 0.5 mg; iron, 75 mg; and selenium, 0.2 mg | |
Statistical analysis: All statistical analyses were performed using the Statistical Analysis System (SAS Institute, 1996). A pen of birds on litter or in metabolism cage constituted the experimental unit. From (3- 6 weeks), 4 dietary treatments were arranged in 6 replications and from (6-7 weeks), 8 treatments were arranged in 3 replications in a randomized complete block design. Means for measurements showing significant differences in the analysis of variance were tested using the PDIFF option. Means±Standard error of the mean (SEM) are presented in the tables and differences were considered statistically significant at p<0.05.
RESULTS
Decreasing dietary TP 6.5 to 5.0 g kg-1 from 3 to 6 weeks had no significant effect on feed intake, body weight gain, breast muscle yield and FCR (Table 2). A dietary TP x sex interaction was not detected at 6 weeks (Table 2). Serum P were lower (p<0.01) for birds fed 5.0 g kg-1 TP compared to the other 3 diets from 3 to 6 weeks. Serum P increased abruptly when TP was increased to 5.5 g kg-1 and as dietary TP increased from 5.5 to 6.5 g kg-1 serum P was not responsive.
Increasing the TP content of the diets from 5.0 to 6.5 g kg-1 (diets 1 to 4) or removing dicalcium phosphate completely from the diets (diets 5 to 8) at 7 weeks of age had no significant effects on feed consumption, body weight gain, breast muscle yield and FCR (Table 3). A gender effect was evident at 7 weeks, breast yield was 8.5% higher in males but a diet x gender interaction was not detected.
| Table 2: | Effect of dietary phosphorus on feed consumption, body weight gain, FCR, breast muscle yield and serum phosphorus content of broiler chicks from 3 to 6 weeks |
| a,bTreatments means within columns followed by different superscripts are significantly different (p<0.05), 1Standard error of the mean, 2Feed consumption and body weight gain per bird during 21 days, 3Each mean based on 12 samples (6 females and 6 males) per treatment taken at 6 weeks of age. The following regression equations were found from calculation of the relationship between serum P and dietary P for females and males, Serum P (F) = -69.37+25.06 *P -2.04 P* P Serum P (M) = -0.881+1.206 * P | |
| Table 3: | Effect of dietary phosphorus on feed consumption, body weight change, FCR, breast muscle yield and serum phosphorus content in broiler chicks from 6 to 7 weeks |
| a,b,c,dTreatments means within columns followed by different superscripts are significantly different (p<0.05), 1Standard error of the Mean. 2Feed consumption and body weight gain per bird during 7 d period, 3Each mean based on 6 samples (3 females and 3 males) per treatment taken at 7 weeks of age | |
At 7 weeks, serum P content was affected by treatment (p<0.01) and sex (p<0.01). Birds that received supplemented diets (diets 1 to 4) had higher serum P compared to the other group (diets 5 to 8), (7.52 vs. 5.07 mg dL-1, respectively). Diets (1 to 4) showed no significant differences among them (p>0.05).
| Table 4: | Effect of dietary phosphorus on broiler chicks Femora and Tibiae measurements at 6 weeks of age |
| a,bTreatments means within columns followed by
different superscripts are significantly different (p<0.05), 1Standard
error of the Mean. 2Each mean based on 12 samples (6 females and 6 males) per treatment taken at 6 weeks of age |
|
On the other hand, serum from females contained 10.4% higher P than males (p<0.05).
Results of bone measurement at 6 weeks are shown in Table 4. Dietary P did not affect femora weights, lengths and diameters (Table 4). Sex had a significant effect on femora weights, lengths and widths (p<0.01), males had heavier, longer and wider femora bones compared to females. Ash content of the femora was significantly influenced by TP and sex (p<0.01). Femora ash content was not consistently responsive to TP level, ash contents were lower for birds receiving 5.0 g kg-1 TP compared to the three other treatments (Table 5). Femora of females contained 3.6% higher ash than males (p<0.05).
Increasing TP content of the diets had no significant effect on tibiae weights, lengths and widths (p>0.05), while sex had a significant effect. Males had heavier, longer and wider tibiae compared to the females (p<0.01). Results of bone ash content are presented in Table 5. Tibiae ash differed between diets and sex (p<0.01); there was an increase in tibia ash content as the level of TP increased in the diets (p<0.01). Birds that received the highest TP had 12% higher tibia ash compared to those that received the lowest TP level. Tibia ash was 6.4% higher in females compared to the results for males.
Dietary P affected toe ash (p<0.01) as documented in Table 5. Birds that received 5.0 and 5.5 g kg-1 TP diets had lower toe ash content compared to the other two groups. Cross-sectional cortical area (A), area moment of inertia (I), polar moment of inertia (J) and diaphysial curvature (θ) were not significantly affected by diet (Table 5). However, I, J and θ were affected by sex (p<0.01, 0.01, 0.05, respectively). Males had 47.8 and 37% higher I and J values, respectively compared to females; also femora from females showed more curvature than femora of males (165 vs. 163), respectively (Table 5).
The bone measurements at 7 weeks showed that femora were affected by sex, males had heavier, longer and wider (p<0.01) femora compared to the females, while diet had no significant effect (Data not shown). Significant effects of sex on tibiae weights, lengths and widths were detected (p<0.01).
| Table 5: | Effect of dietary phosphorus on femur ash content, femur geometric values, tibia ash content and toe ash of broiler chicks at 6 weeks |
| a,b,cTreatments means within columns followed by different superscripts are significantly different (p<0.05), 2Each mean based on 12 samples (6 females and 6 males) per treatment taken at 6 weeks of age, I, A and J were measured at three points, (1/4, ½ and 3/4 of femoral length), the mean based on the average values of the three measurements, 3Area moment of inertia, 4Cross sectional cortical area, 5Polar moment of inertia, 6Diaphyseal curvature, 7Standard error of the mean. The following regression equations were found from calculation of the relationship between tibia ash and dietary P for females and males: Tibia ash (F) = 23.78+3.44 * P Tibia ash (M) = 24.22+2.91 | |
Males showed higher values for the three responses. In contrast, diet had no effect on these responses (p>0.05). Differences in I, A and J were affected by the sex of the bird rather than treatment; males had higher values. Diaphyseal curvature was not affected by treatments or sex of the bird.
Ash content of femora were higher in the inorganic P supplemented compared to unsupplemented group (40.4 vs. 37.3%), respectively. Males had 6.6% lower femora ash compared to the females (p<0.01). Dietary P or gender had no effect on toe ash (p>0.05).Tibiae ash showed an effect due to different dietary treatments (p<0.01). Tibiae from birds fed the inorganic P supplemented diets had 5.9% higher ash content compared to the unsupplemented group. Males had 7.2% lower tibia ash compared to the females (p<0.01).
The effects of dietary treatments on P and Ca retention and P intake and excretion at 6 weeks are shown in Table 6. Birds given 5.5 g kg-1 TP diets retained more P than those given 5.0, 6.0 and 6.5 g kg-1, respectively ((11.5, 46.8 and 43.6%), p<0.01). This group also retained more Ca than those that received 5.0, 6.0 and 6.5 g kg-1 TP, respectively (31.8, 22.0 and 58.9%, p<0.01). Dietary phosphorus level linearly increased P intake and excretion (p<0.01). Birds fed 6.5 g kg-1 TP consumed 13, 22 and 31% and excreted 8.0, 20.5 and 26.5% more P than those fed 6.0, 5.5 and 5.0 g kg-1 TP.
Phosphorus intake and excretion data at 7 weeks are shown in Table 7. Birds that received diets 3 and 4 consumed 56.8 and 49.8% and excreted 25.4 and 31.1% more P than those that received diets 7 and 8, respectively (p<0.01). Birds fed 5.0 g kg-1 TP during growing period and 3.8 g kg-1 TP during finishing period showed a great reduction in P intake and excretion (69.9 and 62.5%, respectively) compared to those fed 6.5 g kg-1 TP during growing and finishing periods (p<0.01).
| Table 6: | Effect of dietary phosphorus on calcium and phosphorus retention and phosphorus excretion at 6 weeks |
| a,b,c Treatments means within columns followed by different superscripts are significantly different (p<0.05), 1Standard error of the mean, 2Based on fecal collection from 6 pens per treatment (2 birds per pen) for 24 h period during week five, 3Based on pooled samples from 4 fecal collections from 6 pens per treatment (2 birds per pen). The following regression equation was found from calculation of the relationship between P intake and dietary P: P intake = -15.83+158.8 * P. The following regression equation was found from calculation of the relationship between P excretion and dietary P: P excretion = 21.9+57.1* P | |
| Table 7: | Effect of dietary phosphorus on P intake and excretion of broiler chicks from 6 to 7 weeks |
| a,b,c,dMeans within columns followed by different superscripts are significantly different (p<0.05), 1Standard error of the mean, 2Each mean based on the pooled collection from 3 pens per treatment taken at 7 weeks of age. The following regression equation was found from calculation of the relationship between P intake and dietary P for treatments 1 to 4: P intake = -8391.8+3113.1 * P - 256.3 P *P. The following regression equation was found from calculation of the relationship between P excretion and dietary P for treatments 1 to 4: P excretion = -226.8+114.1* P | |
DISCUSSION
The results obtained from this study indicated that it is possible to reduce dietary P and as a result reduce P excretion with no adverse effect on the performance from 3 to 7 weeks.
During the growing period, birds which had received diet with 5.0 g kg-1 TP gained weight similarly to those fed the highest level 6.5 g kg-1. This result agreed with the result obtained when breast muscle yield was measured, no significant effects were detected for the dietary treatments. Moreover, TP did not affect feed consumption or FCR. In this experiment, 6.3% reduction in body weight gain from 3 to 6 weeks period was found when the birds were fed 30% less TP but this was not a significant difference. These results are in agreement with those of Skinner et al. (1992), Chen and Moran Jr. (1995), Waldroup et al. (2000) and Dhandu and Angel (2003), who reported that 5.0 g kg-1 P level was required for maximum growth. In agreement with the results obtained by Chen and Moran Jr. (1995) FCR was not affected by the level of P in the diet. Yan et al. (2000) concluded that body weight gain and FCR of broilers from 3 to 6 weeks were similar when birds were fed diet containing 2.6 g kg-1 npP compared to those fed the NRC-recommended level of 3.5 g kg-1. Later, Yan et al. (2001) did not find any significant difference in body weight gain of broilers which had fed levels of npP from 1.5 to 4.5 g kg-1 from 3 to 6 weeks. Recently, Shaw et al. (2010) found that body weight increased significantly with an increase in npP from 3.5 to 5.0 g kg-1 at 4 weeks.
Omitting inorganic P source from the diet during the finishing period decreased TP content of excreta to 71% compared to a 6.5 g kg-1 TP diet. Live performance was not affected when the inorganic P source was omitted from the finishing diets. No significant differences could be detected for any dietary treatment on feed intake, body weight gain and FCR; birds received diets supplemented with inorganic P gained weight similar to those fed diets without inorganic P supplementation. Previous investigators had removed inorganic P source from the withdrawal feed of broiler males without adverse effects on body weight gain, feed intake, feed utilization and carcass yield (Skinner et al., 1992; Chen and Moran Jr., 1994, 1995). Yan et al. (2000) reported that males fed diets with 0.075% less npP did not differ significantly in body weight (at 6 and 8 weeks old) from those fed NRC the level but they detected significant differences when the birds fed diets with 0.15% less than the NRC level. Dhandu and Angel (2003) reported that body weight gain, feed intake and FCR of birds fed 1.0 g kg-1 npP was not different from that of those fed the NRC (1994) level of 3.1 g kg-1 from 6 to 7 weeks.
Serum P showed treatment and sex effects during growing and finishing periods. Gardiner (1962) concluded that the level of plasma inorganic P responded very quickly to suboptimal dietary P levels and to feed withdrawal. During the growing period, birds that received 5.5 g kg-1 TP showed no significant differences than those that received the highest level of TP. During the finishing period, serum P was inconsistently responsive to dietary P levels. Birds that received any level of inorganic P supplementation showed no significant differences among them. Females in both periods had a higher serum P compared to males; this could be explained by the fact that males have more phosphate deposited in their bones since they have longer and heavier bones.
The weight, length and width of femora or tibiae showed no significant differences during growing or finishing periods and were inconsistently responsive to TP levels. Similar results were reported by Skinner et al. (1992). As expected, there was a significant sex effect for the criteria measured and same results were reported by Bond et al. (1991) who reported an increase in bone weight and length for male chicks. Similarly, Applegate and Lilburn (2002) reported a significant sex effect in tibia and femur weight from 28 through 43 days of age.
The skeletal status of chicken is assessed by estimation of tibia ash and occasionally supplemented by estimation of ash composition. The results obtained in this experiment suggested that incremental additions of P during growing and finishing periods improved tibia and femur ash but did not improve the mechanical properties of femur. Previous studies have produced results that are consistent with observations in the current study. Bond et al. (1991) reported similar results for tibia ash weight in females and males at 6 and 7 weeks. In agreement with the results of Applegate and Lilburn (2002), ash percentage was greater in tibia than femur. However, bone ash values in this study were lower than have been observed for chicks of this age receiving the same level of P (Ravindran et al., 1995; Yan et al., 2000). This finding supports the idea that even with the lower level of P the strength and mobility of bones were not affected. Dhandu and Angel (2003) reported that tibia ash weight and percentage of broilers fed 1.5 g kg-1 npP were significantly lower than that of broilers fed 1.9, 2.3 and 3.1 g kg-1 npP. Generally optimum P level for maximum growth is less than that required for maximum bone ash; many investigators have utilized maximum bone ash as the criteria for determining P requirements by chickens. Thorp and Waddington (1997) reported that the inadequacies in cortical bone of chicken would contribute to increased incidence in bone fracture. In this experiment differences in the cortical area between different treatments were not found. There were no problems in mortality or bone fractures during the entire study even when the inorganic P was removed completely from the diets during finishing period. Females were found to have higher percentage of ash. Bond et al. (1991) reported that weight of tibiae ash in males was greater than in females but females had higher percent of ash.
The amount of P in the diet and the efficiency with which the chick utilizes this dietary P are the major factors in determining the amount of P in the excreta. Reducing TP content of the diets markedly reduced fecal P. In this study there were significant reductions in TP intake and excretion from 3 to 7 weeks as a result of P in the diet. Birds that received the lower TP diet consumed and excreted less than those that received higher levels. This agrees with the results obtained by Powell et al. (2008) who reported that TP was decreased in the litter of broilers by 21.5% in the low P diet (a 0.05% reduction in npP compared to the control).
Increased dietary TP resulted in reduced P and Ca retention is in agreement with the results of Yi et al. (1996) who found that retention of P and Ca linearly decreased as the level of the npP increased in the diets. This may be due to the lower intestinal phytase activity and as a result lower PP utilization (Abudabos et al., 2000). Fox et al. (1981) reported that feeding diets low in P increased the duodenal absorption of P.
The results of this study indicate that the TP levels needed by broilers to maintain growth and feed utilization from 3 to 7 weeks are less than those needed for maximum tibia ash. On the other hand, retention of P will decrease as the P consumed surpasses the requirement. Birds utilized PP, especially those that received a low level of inorganic source during the growing period and no supplementation during finishing period. In conclusion, the results of this study clearly show that dietary P could be reduced to 5.0 g kg-1 from 3 to 6 weeks and omitted from the withdrawal diets without adverse effects on live performance or skeletal integrity and this could decrease P intake and as a result P excretion.
ACKNOWLEDGMENT
This project was supported by King Saud University, Deanship of Scientific Research, College of Food and Agriculture Sciences, Research Center.