Effects of Phytase Supplementation on the Performance, Egg Quality, and Phosphorous Excretion of Laying Hens Fed Different Levels of Dietary Calcium and Nonphytate Phosphorous H. S. Lim, H. Namkung, and I. K. Paik1 Department of Animal Science, Chung-Ang University Ansung-Si, Kyonggi-Do 456-756, Korea the second period. Low Ca decreased egg specific gravity, eggshell strength, and eggshell thickness in both periods and increased Haugh units in the second 10-wk period. Phytase supplementation decreased the percentage of broken and soft-shell eggs. High NPP increased fiber availability but decreased Ca availability. High Ca decreased Ca availability, whereas phytase increased availability of dry matter, fiber, and P. High NPP increased retention of P and Fe but also increased excretion of P. High Ca decreased retention of Zn and Fe. Phytase supplementation increased P retention, resulting in decrease of P excretion. In conclusion, supplementation of microbial phytase at a level of 300 U per kg diet of laying hens can improve egg production, decrease broken and soft egg production rate, and P excretion. The effects of phytase supplementation are significantly modified by the level of Ca and NPP.
ABSTRACT An experiment employing a factorial arrangement of two levels (3.0 and 4.0%) of Ca, two levels (0.15 and 0.25%) of nonphytate phosphorus (NPP), and two levels (0 and 300 U/kg diet) of microbial phytase was carried out with 960 ISA-brown layers from 21 to 41 wk of age. There was a significant interaction between NPP level and phytase for egg production. High NPP level and phytase supplementation increased egg production only in the second 10-wk period (31 to 41 wk). High NPP and low Ca increased feed intake, and a significant interaction between levels of NPP and Ca was observed in the first 10 wk. High NPP improved feed efficiency only in the second 10-wk period. Low NPP improved egg specific gravity and eggshell thickness but decreased Haugh units in the first 10-wk period; high NPP decreased the percentage of broken and soft-shell eggs in
(Key words: layer, microbial phytase, nonphytate P, Ca, laying performance, eggshell quality) 2003 Poultry Science 82:92–99
Krikorian, 1982). Phytic acid can form insoluble salts with Ca, Mg, Fe, Zn, Cu, and Mn (Oberleas, 1973; Morris, 1986; Bedford and Schulze, 1998; Liu et al., 1998). When phytic acid is hydrolyzed by microbial phytase, it may release all phytate-bound minerals such as Ca, P, Mg, Cu, Zn, Fe, and K (Sebastian et al., 1996). Phytase also improved N absorption in laying hens (Van der Klis and Versteegh, 1991) and improved nitrogen and amino acid digestibilities in broilers (Ravindran et al., 2001). The mineral level of the diet can also affect the degree of phytate degradation (Sandberg et al., 1993). High levels of dietary Mg and Ca are known to decrease intestinal phytase activities in chicks (McCuaig et al., 1972). Supplemental microbial phytase in a corn-soybean meal diet improves phytate P utilization more effectively at moderately low levels of dietary Ca than at normally recommended levels for pigs (Lei et al., 1994). Qian et al., (1996) reported that widening the Cato-total-P ratio in the diet from 1.4 to 2.0 decreased the phytase efficacy in the performance of turkey. There are
INTRODUCTION Approximately two-thirds of the P in plant feedstuffs is present as phytic acid in the form of myo-inositol hexaphosphates (Cromwell, 1980). Phytate P has low P availability, which leads to the use of an inorganic P source to meet the P requirement of most monogastric animals such as poultry and pigs (Peeler, 1972). Peter (1992) reported that laying hens fed a low nonphytate P (NPP) diet with phytase had significantly higher egg production, egg weights, and feed consumption than hens consuming the low NPP diet without supplemental phytase. Negative influences of phytic acid on the solubility of proteins and the function of pepsins can be expected because of the ionic binding between basic phosphate groups of phytic acid and protonized amino acids, such as lysyl, histidyl, and arginyl residues (Camus and Laporte, 1976; Singh and
2003 Poultry Science Association, Inc. Received for publication April 4, 2002. Accepted for publication August 7, 2002. 1 To whom correspondence should be addressed:
[email protected].
Abbreviation Key: NPP = nonphytate phosphorus; TCP = tricalcium phosphate.
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PHYTASE, NONPHYTATE PHOSPHOROUS, AND CALCIUM IN EGG PRODUCTION TABLE 1. Diet composition 3% Ca 0.15% NPP1
Ingredients Corn Soybean meal-44 Wheat bran Corn gluten Rapeseed meal Animal fat Tricalcium phosphate (19% P, 32% Ca) Limestone (37% Ca) NaCl DL-Methionine - 50% Layer premix2 Phytase3 (300 U/g) Sand Total Calculated composition ME, cal/kg CP, % Arg, % Lys, % Met, % Met and Cys, % Ca, % NPP, % Total P, % Salt, % Analyzed value Ca, % Total P, %
4% Ca 0.25% NPP
0.15% NPP
0.25% NPP
59.81 15.77 5.60 3.26 3.00 1.50 0.13 7.65 0.31 0.11 0.10 – 2.77 100
59.81 15.77 5.60 3.26 3.00 1.50 0.13 7.65 0.31 0.11 0.10 + 2.77 100
59.81 15.77 5.60 3.26 3.00 1.50 0.64 7.20 0.31 0.11 0.10 – 2.70 100
59.81 15.77 5.60 3.26 3.00 1.50 0.64 7.20 0.31 0.11 0.10 + 2.70 100
59.81 15.77 5.60 3.26 3.00 1.50 0.13 10.35 0.31 0.11 0.10 – 0.07 100
59.81 15.77 5.60 3.26 3.00 1.50 0.13 10.35 0.31 0.11 0.10 + 0.07 100
59.81 15.77 5.60 3.26 3.00 1.50 0.64 9.91 0.31 0.11 0.10 – 0 100
59.81 15.77 5.60 3.26 3.00 1.50 0.64 9.91 0.31 0.11 0.10 + 0 100
2,750 16.00 0.98 0.70 0.35 0.61 3.00 0.15 0.41 0.37
2,750 16.00 0.98 0.70 0.35 0.61 3.00 0.15 0.41 0.37
2,750 16.00 0.98 0.70 0.35 0.61 3.00 0.25 0.51 0.37
2,750 16.00 0.98 0.70 0.35 0.61 3.00 0.25 0.51 0.37
2,750 16.00 0.98 0.70 0.35 0.61 4.00 0.15 0.41 0.37
2,750 16.00 0.98 0.70 0.35 0.61 4.00 0.15 0.41 0.37
2,750 16.00 0.98 0.70 0.35 0.61 4.00 0.25 0.51 0.37
2,750 16.00 0.98 0.70 0.35 0.61 4.00 0.25 0.51 0.37
3.12 0.43
3.14 0.43
3.15 0.50
3.17 0.51
3.99 0.42
3.96 0.40
4.05 0.49
3.97 0.51
1 Provides per kg: vitamin A, 8,800,000 IU; vitamin D3, 2,500,000 IU; vitamin E, 5,000 IU; vitamin K3, 500 mg; vitamin B1, 800 IU; vitamin B2, 2,200 mg; vitamin B6, 3,000 mg; vitamin B12, 3,000 µ mg; niacin, 10,000 mg; folic acid, 250 mg; Zn, 53,000 mg; Mn, 70,000 mg; Fe, 8000 mg; Cu, 5,000 mg; 1,850 mg; Co, 250 mg; Mo, 100 mg; Se, 100 mg. 2 Novo Nordisk Corp., Novo Alle, 2880 Bagsvaerd, Denmark, phytase activity.
inconsistencies in the literature regarding the effect of phytase on eggshell quality. Several investigators reported a beneficial effect on eggshell quality from phytase supplementation (Gordon and Roland, 1997; Punna and Roland, 1999), whereas others did not observe any beneficial effect (Van der Klis et al., 1997; Parsons, 1999). The experiment reported here was conducted to determine the effects of a microbial phytase product on the productivity, eggshell quality and P excretion of laying hens (ISA brown) fed different levels of dietary Ca and NPP.
MATERIALS AND METHODS Experimental Diet Diets were formulated to have the same nutrient density except for Ca and P (Table 1). Eight experimental diets were factorially arranged as two levels of Ca (3 and 4%), two levels of NPP (0.15 and 0.25%), and two levels of microbial phytase (0 and 300 U/kg diet). The microbial phytase was a product of Novo Nordisk Corporation,2 which is made from a genetically modified Aspergillus oryzae and contains 2500 unit/g. Tricalcium phosphate (TCP) was used in diets as a source of NPP.
2
Novo Nordisk Corp., Novo Alle, 2880, Bagsvaerd, Denmark.
Feeding Regimen Nine hundred and sixty 21-wk-old ISA brown pullets were assigned to eight dietary treatments. Each treatment consisted of four replications of 15 cages (two birds per cage). Diets were presented in mash form, and feed and water were given ad libitum during the experimental periods. Birds were provided with programmed lighting and ventilation. The lighting program started with 14:30 h of light on the initiation of the experiment and then increased 30 min every wk until 16:00 h of light was achieved.
Parameters of Production Performance and Egg Quality The number of total eggs, broken and soft-shell eggs and egg weight was determined on a daily basis from 21 to 41 wk of age. Feed consumption was measured weekly. Egg quality was measured starting from 50% average henday egg production (age = 23 wk). Sixty eggs from each treatment (15 eggs per replication) were collected randomly every week to measure egg specific gravity. Specific gravity of eggs was determined by using salt solutions made of incremental concentration of 0.005 in the range from 1.070 to 1.110. After measuring specific gravity, 30 eggs in the medium specific gravity range (from 1.085 to 1.095) from each treatment were sampled to measure shell strength, shell thickness, and Haugh units. Shell
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LIM ET AL.
strength was measured by using the Compression Test Cell in Texture Test Systems.3 Shell thickness was a mean value of measurements at three locations on the egg (air cell, equator, and sharp end) measured by using dial pipe gauge.4 Haugh units were calculated with the HU formula (Eisen et al., 1962), based on the height of albumen determined by a micrometer5 and egg weight.
Metabolic Trial After 30 wk on experimental diets, five birds per treatment (at least one hen from each replication) kept for metabolic trials were individually housed in metabolic cages. After 4 d of adjustment, excreta were collected for 3 d. Diets and excreta were analyzed for proximate components by AOAC (1990) method. Mineral contents were assayed by using an inductively coupled plasma emission spectrometer.6
Statistical Analysis Data were subjected to ANOVA using general linear model procedure (SAS Institute, 1990) after adjusting for age effects. Data were expressed in means of two separate periods (21 to 30 wk and 31 to 41 wk) to represent the change of egg production rate. Significant differences among treatment means were measured by Duncan’s multiple range test at P < 0.05.
RESULTS Production Performance Egg production, egg weight, feed intake, and feed conversion are shown in Table 2. Egg production in the first 10-wk period was not affected by dietary treatments. In the second 10-wk period, a NPP level by phytase interaction (P < 0.05) was observed, indicating that the effect of phytase on egg production was more pronounced in hens fed 0.25% NPP than in hens fed 0.15% NPP. However, hens fed 0.25% NPP and phytase showed significantly greater egg production. Ca level did not affect egg production. Egg weight was not significantly affected by sources, but it followed a general tendency of egg weight being inversely proportional to egg production. Feed intake in the first period was significantly greater with 0.25% NPP and 3.0% Ca diets than with 0.15% NPP and 4.0% Ca diets, respectively, and interaction of the level of NPP and Ca was also significant in the first period. Feed conversion was better (P < 0.05) for birds fed 0.25% NPP diets than for those fed 0.15% NPP diets.
Egg Quality Specific gravity tended to decline as the birds got older (Table 3). There were significant effects of the levels of 3
Model Model 5 Model 6 Model 4
T2100C, Food Technology Corp., Rockville, MD. 7360, Mitutoyo Corp., Kawasaki, Japan. S-8400, AMES, Waltham, MA. JY-24, Jobin Yvon, Longjumeau, Cedex, France.
NPP and Ca on egg specific gravity in both periods. Egg specific gravity was greater in hens fed the 0.15% NPP diets and 4.0% Ca diets than those fed 0.25% NPP diets and 3.0% Ca diets. Phytase alone did not affect egg specific gravity. The three-way interaction of phytase, NPP, and Ca on egg specific gravity was observed in both periods. Specific gravity was highest in low NPP and high Ca treatments regardless of phytase supplementation. Phytase supplementation increased specific gravity except in the high NPP and low Ca treatments, in which phytase supplementation decreased specific gravity of egg. Haugh units of eggs were high in hens fed 0.25% NPP diets in the first period but high in hens fed 3.0% Ca diets in the second period. The Ca level by phytase interaction was significant for Haugh units in the second period. The 4.0% Ca diet increased eggshell strength in both periods. The Ca by NPP interaction was significant in the first period, in which the highest shell strength was observed in the treatment with high levels of NPP and Ca. The NPP levels by phytase interaction were significant in the second period. Shell thickness increased in hens fed 0.15% NPP or 4.0% Ca in the first period, but by only 4.0% Ca in the second period. Phytase supplementation decreased broken and soft egg production in both periods. The 0.25% NPP diet treatments showed lower broken and soft egg production in the second period. A three-way interaction of phytase, NPP, and Ca was observed; phytase supplementation to the diets with 0.15% NPP and 3.0% Ca or diets with 0.25% NPP and 4.0% Ca decreased broken and soft egg production.
Availability, Retention, and Excretion of Nutrients The availability of nutrients is presented in Table 4. Analysis of main effects indicated that 0.25% NPP increased availability of fiber, 0.25% NPP and 4.0% Ca decreased availability of Ca, and phytase increased availability of DM, fiber, and phosphorus. The two-way interaction of NPP and Ca on the availability of fat and Mg was observed; at 0.25% NPP, 4.0% Ca decreased fat and Mg availability. Interaction of NPP by phytase on the availability of Zn was significant; supplementation of phytase to the 0.25% NPP and 4% Ca diet increased the availability of fiber. Interaction of phytase and Ca on Zn availability was observed; phytase supplementation to the 4.0% Ca diets increased Zn availability, whereas phytase supplementation to the 3.0% Ca diets decreased Zn availability. The retention and excretion of nutrients expressed on daily intake basis are shown in Table 5. The 0.25% NPP diets increased the retention of P and Fe and excretion of P. The 4.0% Ca diets decreased the retention of Zn and Fe but increased Ca excretion. Phytase supplementation increased P retention and decreased P excretion. Interactions of NPP and Ca for retention of Mg and Zn and excretion of Zn were significant. Mg and Zn retention was lower in hens fed 4.0% Ca diet when NPP level was 0.25%. Zn excretion was lower in birds fed 0.15% NPP diet with 3.0% Ca but high in birds fed 0.25% NPP diet with 4.0% Ca. The interaction of NPP and phytase on Zn retention
3 3 4 4 3 3 4 4
0.15 0.15 0.15 0.15 0.25 0.25 0.25 0.25 SEM Main effect NPP, % 72.56 75.06 75.62 72.00 73.21 74.41
0.15 0.25 3.0 4.0 0 300 0.3745 0.1988 0.6693 0.6159 0.7058 0.8759 0.9700
75.27 74.87 69.71 70.39 75.20 77.12 72.66 75.25 8.8696
21 to 30 wk
0 300 0 300 0 300 0 300
Phytase (U/kg diet)
a-d
0.8118 0.0840 0.3007 0.6351 0.1325 0.2112 0.7831
54.99 55.07 55.31 54.75 55.20 54.86
87.68b 89.44a 89.09 88.03 87.76b 89.36a 0.0019 0.0623 0.0046 0.6014 0.0314 0.3181 0.0846
55.11 55.58 54.72 55.55 55.45 55.12 55.52 54.21 1.0215
21 to 30 wk
0.2713 0.6767 0.0688 0.0561 0.8254 0.3198 0.8295
59.54 59.30 59.47 59.38 59.61 59.23
59.47 59.29ab 59.96a 59.45ab 59.64ab 59.46ab 59.40ab 58.70b 0.7135
ab
31 to 41 wk
Egg weight (g)
87.39 89.32abc 87.58cd 86.43d 88.62bcd 91.03a 87.45cd 90.68ab 1.8684
cd
31 to 41 wk
Hen-day egg production (%)
Means within each column with no common superscript differ (P < 0.05).
NPP Ca Phytase NPP × Ca NPP × phytase Ca × phytase NPP × Ca × phytase
Phytase, U/kg diet
Ca, %
Ca (%)
NPP (%)
Treatment
P 0.0337 0.0001 0.5148 0.0155 0.3784 0.2586 0.2296
105.62b 106.86a 107.73a 104.74b 106.05 106.43
107.39 108.24a 102.95c 103.88c 107.03ab 108.26a 106.82ab 105.33bc 1.8412
ab
21 to 30 wk
0.4112 0.0612 0.3161 0.7590 0.9636 0.2648 0.9853
114.41 115.14 115.60 113.94 115.22 114.33
115.30 115.45 114.37 112.51 112.81 115.85 115.39 113.49 2.9390
31 to 41 wk
Feed intake (g/day)
0.5300 0.2032 0.7486 0.5016 0.5793 0.8789 0.7655
2.61 2.58 2.56 2.63 2.60 2.59
2.56 2.57 2.65 2.67 2.58 2.55 2.63 2.56 0.0499
21 to 30 wk
0.0495 0.7239 0.1495 0.4399 0.3384 0.7616 0.4052
2.18a 2.13b 2.15 2.16 2.17 2.14
2.21a 2.17ab 2.16ab 2.17ab 2.15ab 2.10ab 2.17ab 2.10b 0.0222
31 to 41 wk
Feed conversion (g/100 g egg mass)
TABLE 2. Effect of phytase supplementation and level of NPP and Ca in the diet on egg production, egg weight, feed intake, and feed conversion
PHYTASE, NONPHYTATE PHOSPHOROUS, AND CALCIUM IN EGG PRODUCTION
95
3 3 4 4 3 3 4 4
0.15 0.15 0.15 0.15 0.25 0.25 0.25 0.25 SEM Main effect NPP, % 1.0925a 1.0917b 1.0913b 1.0927a 1.0920 1.0921
0.15 0.25 3.0 4.0 0 300 0.0101 0.0001 0.1154 0.8762 0.0042 0.5321 0.0026
1.0907 1.0922bc 1.0927ab 1.0932a 1.0915cd 1.0904e 1.0921bc 1.0926ab 0.0020
de
21 to 30 wk
0 300 0 300 0 300 0 300
Phystase (U/kg diet)
0.0001 0.0001 0.3398 0.1647 0.3910 0.0142 0.0001
1.0902a 1.0895b 1.0892b 1.0904a 1.0898 1.0899
1.0892 1.0899bc 1.0910a 1.0908a 1.0896cd 1.0884e 1.0893cd 1.0905ab 0.0020
d
31 to 41 wk
a–e
0.0101 0.2267 0.3276 0.2232 0.0534 0.2069 0.8889
90.74b 91.58a 91.36 90.96 91.00 91.32
90.89 91.38a 89.64b 91.04a 91.92a 91.24a 91.55a 91.61a 2.5269
ab
0.0908 0.0112 0.0791 0.1582 0.5904 0.0332 0.9899
85.65 86.21 86.36a 85.50b 86.22 85.63
86.87 85.75ab 84.83b 85.13b 87.15a 85.66ab 86.05ab 86.00ab 3.0569
a
31 to 41 wk
Haugh unit 21 to 30 wk
Means within each column with no common superscript differ (P < 0.05).
NPP Ca Phytase NPP × Ca NPP × phytase Ca × phytase NPP × Ca × phytase
Phytase, U/kg diet
Ca, %
Ca (%)
NPP (%)
Specific gravity
0.7909 0.0006 0.3673 0.0112 0.3151 0.3357 0.4540
4.501 4.492 4.437b 4.555a 4.481 4.511
4.516 4.454bc 4.489abc 4.544ab 4.360c 4.418bc 4.558ab 4.630a 0.0484
ab
21 to 30 wk
P 0.1085 0.0001 0.9537 0.6238 0.0344 0.9949 0.1593
4.670 4.623 4.586b 4.707a 4.646 4.647
4.627 4.607c 4.775a 4.672abc 4.543c 4.566c 4.637bc 4.744ab 0.0421
bc
31 to 41 wk
Eggshell strength (kg/Egg)
0.0017 0.0070 0.2386 0.0987 0.6315 0.3182 0.1238
0.360a 0.356b 0.357b 0.360a 0.359 0.357
0.358 0.357bc 0.363a 0.361ab 0.358bc 0.354c 0.356c 0.358bc 0.0085
bc
21 to 30 wk
0.0679 0.0002 0.1985 0.0507 0.9768 0.7114 0.0277
0.379 0.378 0.377b 0.380a 0.379 0.378
0.376 0.377bc 0.384a 0.380ab 0.378bc 0.375c 0.378bc 0.379bc 0.0083
bc
31 to 41 wk
Eggshell thickness (mm)
TABLE 3. Effect of phytase supplementation and level of NPP and Ca in the diet on egg and egg shell characteristics
0.2307 0.8300 0.0222 0.7193 0.4565 0.6323 0.0029
1.11 0.92 1.03 1.00 1.20a 0.83b
1.60 0.71bc 1.11abc 1.06abc 0.75bc 1.06abc 1.33ab 0.53c 0.5032
a
21 to 30 wk
0.0158 0.8493 0.0006 0.1440 0.9764 0.5163 0.0001
1.09a 0.83b 0.97 0.95 1.14a 0.77b
1.57a 0.78cd 0.97bc 1.02bc 0.67cd 0.85cd 1.36ab 0.43d 0.3567
31 to 41 wk
Broken & soft egg production (%)
96 LIM ET AL.
97
PHYTASE, NONPHYTATE PHOSPHOROUS, AND CALCIUM IN EGG PRODUCTION TABLE 4. Effect of phytase supplementation and level of NPP and Ca on nutrient availability of the diets Treatment NPP (%) 0.15 0.15 0.15 0.15 0.25 0.25 0.25 0.25 SEM Main effect means NPP, % Ca, % Phytase, U/kg diet
NPP Ca Phytase NPP × Ca NPP × phytase Ca × phytase NPP × Ca × phytase
Nutrients (%) Ca (%) 3 3 4 4 3 3 4 4
Phytase (U/kg)
DM
Protein
Fat c
Ash
Fiber b
NFE
Ca a
P bcd
Mg
Zn
Fe
0 300 0 300 0 300 0 300
78.2 81.8 78.7 81.5 76.8 84.0 83.3 82.3 1.53
66.6 65.5 65.7 68.2 63.7 68.2 68.2 63.1 2.47
84.0 91.0a 90.4ab 90.3ab 89.4ab 89.9ab 85.5bc 85.8abc 1.28
56.8 52.5 56.8 56.7 43.1 62.9 50.4 47.5 4.51
8.4 9.1b 16.8ab 16.4ab 15.2ab 25.6a 11.5b 26.1a 3.13
86.5 86.6 88.3 86.1 87.0 87.6 87.2 88.6 0.85
82.2 75.6ab 65.2bc 67.5bc 64.8bc 74.6ab 57.8c 61.4bc 3.24
36.6 43.3abc 34.4cd 46.3a 35.1cd 48.2a 33.5d 44.1ab 2.12
28.2 23.9 33.4 27.8 28.0 38.0 24.8 18.9 2.98
49.2 43.8 42.1 46.3 52.5 44.6 36.5 52.5 2.93
50.4 39.9 38.7 36.4 39.0 45.7 41.2 38.3 2.98
0.15 0.25 3.0 4.0 0 300
80.06 81.61 81.48 80.19 79.24b 82.43a
66.38 65.81 66.01 66.18 66.06 66.13
88.90 87.64 88.56 87.99 87.33 89.22
55.69 50.94 53.81 52.82 51.76 54.88
12.64b 19.59a 14.56 17.63 12.96b 19.27a
86.84 87.59 86.91 87.53 87.24 87.19
72.6a 64.6b 72.6a 64.6b 67.5 69.7
40.2 40.2 40.2 40.2 34.9b 45.5a
28.3 27.4 28.3 27.4 28.6 27.1
45.3 46.1 45.3 46.1 45.1 46.4
38.8 41.0 41.2 38.6 39.8 40.1
0.2649 0.3527 0.0262 0.4093 0.1055 0.9844 0.1847
0.7988 0.9419 0.9752 0.8186 0.4654 0.8803 0.1638
0.2771 0.6208 0.1084 0.0044 0.1260 0.1991 0.1447
0.2472 0.8072 0.4444 0.4545 0.2620 0.1961 0.1048
0.0186 0.2746 0.0312 0.0999 0.7809 0.0356 0.6401
P 0.3310 0.4209 0.9530 0.9697 0.6071 0.1901 0.3098
0.0214 0.0019 0.4910 0.7099 0.8428 0.1832 0.2535
0.5606 0.5606 0.0001 0.4547 0.7477 0.5629 0.3717
0.7655 0.2754 0.6193 0.0144 0.1633 0.2502 0.2313
0.7926 0.2330 0.6536 0.5198 0.0119 0.5198 0.2851
0.4705 0.3893 0.9387 0.9947 0.3481 0.5802 0.5218
Means within each column with no common superscript differ (P < 0.05).
a–e
was significant; phytase supplementation to 4.0% Ca diets increased Zn retention. There was a three-way interaction for Ca retention: Ca retention increased when phytase was supplemented to 0.25% NPP diet with 3.0% Ca but decreased when supplemented to 0.25% NPP diet with 4.0% Ca.
DISCUSSION Our findings that high NPP level and phytase increased egg production in the second 10-wk period (31 to 40 wk) agreed with those of Scott et al. (1999), who reported increased egg production in hens fed a phytase-supplemented diet or high-NPP diet from 36 to 51 wk. The interaction between phytase and NPP levels showed that phytase increased egg production in hens fed a diet containing 0.25% NPP, but not in hens fed the diet with 0.15% NPP and 4.0% Ca. Gordon and Roland (1997), however, reported in a study employing a factorial arrangement of NPP levels and phytase that feeding a 0.1% NPP without supplemental phytase decreased egg production, but 0.1% NPP diet supplemented with phytase completely corrected the adverse effect. Boling et al. (2000) obtained similar results in a study involving 0.15% NPP with phytase. In the present experiment, which involved three dietary factors of phytase, NPP, and Ca, there were two-way interactions of phytase and NPP level and also a three-way interaction (P < 0.08). The effect of phytase supplementation on egg production appears to be affected not only by NPP level but also by Ca levels because egg production was significantly greater in birds fed phytase, 0.15% NPP, and 3.0% Ca diet than those fed phytase, 0.15% NPP, and 4.0% Ca diet.
Low dietary Ca level was associated with increased feed intake and reduced eggshell quality. These results are in agreement with a previous study (Roland et al., 1985). Egg specific gravity was increased in hens fed high Ca and low NPP. Gordon and Roland (1998) reported increased egg specific gravity in hens fed high Ca diet, while Keshavarz (2000) reported increased egg specific gravity in low NPP diet. Nahashon et al. (1994) also reported that egg specific gravity was higher in hens fed 0.25% than those on 0.45% NPP diet. Egg specific gravity tended to decrease in the second 10-wk period compared with that of the first period. Gordon and Roland (1997) reported that egg specific gravity decreased over a time period of 2 to 16 wk. Haugh units increased in eggs from hens fed 0.25% NPP diets but decreased in eggs from hens fed 4.0% Ca diets. The reason for the difference was not explainable, but the results correlated with feed intake of treatments. Specific gravity and first-period eggshell thickness were greater with low NPP, whereas percentage of broken and soft-shell eggs were higher with 0.15 than with 0.25% NPP in the second period. This result seems contradictory. However, eggshell strength was not affected by NPP level. Undetermined factors other than specific gravity and eggshell thickness may have influenced eggshell strength and percentage of broken and soft-shell eggs. As expected, eggshell strength and thickness were higher in eggs from hens fed 4.0% Ca diets than in those fed the 3.0% Ca diet. Percentage of broken and soft-shell eggs was decreased by phytase supplementation in both periods and in high NPP diets in the second period. Dietary phytase did not have any consistent effect on eggshell quality. Some investigators observed a beneficial effect on eggshell quality of phytase supplemen-
0.15
4
3
2.05 1.99 2.07 1.97 1.98 2.06
0.15 2.50 3.0 4.0 0 300 0.6474 0.4429 0.5282 0.4219 0.3589 0.7824 0.2861
2.03 2.06 2.02 2.09 1.90 2.30 1.95 1.80 0.22
N
0 300 0 300 0 300 0 300 0.84
Phytase (U/kg)
0.6438 0.2685 0.6936 0.1193 0.0668 0.4343 0.0654
8.75 8.40 9.00 8.15 8.43 8.73
8.73 8.42 9.07 8.79 7.55 11.30 8.36 6.40 0.02
Ash
0.1446 0.3138 0.6636 0.9655 0.2080 0.8248 0.0281
2.93 2.64 2.88 2.68 2.74 2.83
3.11 2.96 2.71 2.95 2.32 3.15 2.83 2.26 0.01
Ca
a–d
0.0290 0.2615 0.0130 0.5716 0.5716 0.3113 0.2177
0.4093 0.2189 0.6787 0.0063 0.1042 0.2189 0.4093
0.05 0.06 0.06 0.05 0.06 0.05
0.19b 0.23a 0.22 0.20 0.18b 0.23a
b
Mg 0.05 0.04b 0.07ab 0.05b 0.06ab 0.09a 0.05b 0.04b 1.15
b
0.18 0.20b 0.16b 0.20b 0.19b 0.29a 0.19b 0.23ab 0.38
P
Retention (g/hen/day)
Means within each column with no common superscript differ (P < 0.05).
NPP Ca Phytase NPP × Ca NPP × phytase Ca × phytase NPP × Ca × phytase
Phytase, U/kg
Ca, %
3
0.15 0.15 0.15 0.15 0.25 0.25 0.25 0.25 SEM Main effect means NPP, %
4
Ca (%)
NPP (%)
Treatment
0.2399 0.0205 0.2711 0.0167 0.0191 0.2877 0.9910
3.54 4.00 4.24a 3.29b 3.55 3.98
3.99 3.05bc 3.07bc 4.04ab 5.02a 4.90a 2.13c 3.94ab 0.06
ab
Zn bc
0.0079 0.0046 0.5481 0.9776 0.2611 0.4304 0.2218
12.70b 16.03a 16.16a 12.57b 14.02 14.72
13.76 15.27ab 10.03c 11.76bc 16.54ab 19.08a 15.74ab 12.76bc 0.78
Fe
P 0.8546 0.3624 0.4192 0.9235 0.7601 0.6317 0.1162
1.02 1.01 1.04 0.99 0.99 1.04
1.00 1.08 1.02 0.97 1.05 1.03 0.89 1.07 0.18
N
0.3623 0.4852 0.3601 0.8717 0.8139 0.2555 0.3675
7.07 7.71 7.64 7.14 7.71 7.07
7.05 7.69 6.92 6.61 9.03 6.77 7.85 7.20 0.01
Ash
0.1597 0.0001 0.4009 0.4988 0.5625 0.7847 0.0866
1.27 1.51 1.03b 1.75a 1.46 1.32
0.92 1.02bc 1.72ab 1.44abc 1.37abc 0.81c 1.83a 2.02a 0.01
c
Ca
cd
0.0001 0.9367 0.0047 0.5264 0.9367 0.6916 0.0877
0.29b 0.34a 0.32 0.32 0.34a 0.30b
0.30 0.29cd 0.31cd 0.26d 0.37a 0.31cd 0.36ab 0.34abc 0.22
P
0.1803 0.1356 0.8105 0.1003 0.4736 0.4736 0.3820
0.15 0.16 0.15 0.16 0.15 0.15
0.15 0.15 0.15 0.15 0.15 0.14 0.16 0.17 1.65
Mg
Excretion (g/hen/day)
TABLE 5. Effect of phytase supplementation and level of NPP and Ca in the diet on retention and excretion of nutrients
0.7172 0.3709 0.9889 0.0091 0.7982 0.4827 0.8069
4.23 4.15 4.09 4.29 4.19 4.19
3.74 3.90 4.56 4.72 4.49 4.23 3.96 3.92
Zn
0.9586 0.1449 0.4334 0.2142 0.5286 0.3755 0.2137
23.91 23.82 22.63 25.11 24.52 23.21
21.01 22.23 26.64 25.76 26.61 20.66 23.84 24.19
Fe
98 LIM ET AL.
PHYTASE, NONPHYTATE PHOSPHOROUS, AND CALCIUM IN EGG PRODUCTION
tation (Gordon and Roland, 1997; Punna and Roland, 1999), but others did not find any effect (Van der Klis et al., 1997; Parsons, 1999). Phytase improved retention of major nutrients and minerals in the current study. These results were similar to a previous study (Um and Paik, 1999) that retention of fiber, fat, and minerals increased after phytase supplementation to a diet containing low levels of NPP. This improved retention may be explained by the fact that phytate complexes were, to some extent, cleaved by phytase (Nair et al., 1991). Increased P retention by phytase resulted in a reduction of P excretion by about 13%. Low Ca diet increased retention of Zn and Fe and decreased Ca excretion. High NPP diet increased retention of P and Fe and increased P excretion. Boling et al. (2000) reported increased P excretion by hens fed a high NNP diet over hens fed a low NPP diet. We concluded that supplementation of microbial phytase at a level of 300 U per kg in the diet of laying hens can improve egg production and decrease the number of broken and soft eggs and P excretion. The effects of phytase supplementation are significantly modified by the levels of Ca and NPP.
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