Evaluation of Phytase Enzyme with Chicks Fed Basal Diets Containing Different Soybean Meal Samples

2006 Poultry Science Association, Inc.

Evaluation of Phytase Enzyme with Chicks Fed Basal Diets Containing Different Soybean Meal Samples M. K. Manangi and C. N. Coon1 The Center of Excellence for Poultry Science, University of Arkansas, Fayetteville 72701

SUMMARY Soybean meal contains approximately 0.62% total P of which 0.4% can be phytate P, which is considered less biologically available for poultry than other forms of P. Soybean meal is a key ingredient in poultry feeds and information is needed about the range of phytate P and nonphytate P in different soybean meals. The phytate P content of soybeans may vary due to climatic conditions, soil type and soybean variety. Previous research has shown that phytate P can be hydrolyzed in the gastrointestinal tract providing available P by adding a commercial phytase enzyme to poultry feed. The extent of phytate hydrolysis by dietary supplementation of phytase has been shown to vary depending on the type of dietary ingredients such as corn, soybean meal, canola meal, and wheat. Research is needed to determine if different commercially available soybean meals respond in a similar manner to a feed added phytase. Twenty-five soybean meal samples were collected from active soybean crushing plants in the United States and 18 of the samples were selected to evaluate the effect of a microbial phytase on phytate P disappearance using 5-d bioassays. The range of analyzed values in soybean meal samples for total P, phytate P, Ca, protein, and neutral detergent fiber (NDF) were 0.59 to 0.87, 0.32 to 0.42, 0.28 to 0.54, 40.44 to 51.69, and 7.78 to 16.09%, respectively. Bioassay results indicate that body weight, feed consumption, and feed conversion ratio improved significantly (P < 0.05) in some of the groups fed diets with enzyme compared with groups fed the same diet with no added enzyme. The range of total P retention and phytate P disappearance for groups fed diets with no enzyme were 21.35 to 48.41 and 13.64 to 37.13%, respectively. The addition of phytase increased total P retention and phytate P disappearance from 56.81 to 68.62 and 76.18 to 94.08%, respectively. The results indicate no correlation among components (total P, phytate P, Ca, protein, and NDF) of soybean meal samples, percentage of phytate P disappearance, and percentage of total P retention for groups fed diets with and without added phytase. Key words: phytate phosphorus disappearance, soybean meal source, broiler 2006 J. Appl. Poult. Res. 15:292–306

DESCRIPTION OF PROBLEM Phytate phosphorus (myo-inositol 1,2,3,4,5,6hexakis dihydrogen phosphate; IP6) is the main P-containing constituent of many seeds and tubers 1

Corresponding author: [email protected]

and is rapidly hydrolyzed by phytases soon after germination or sprouting [1]. Aleurone particles are the major sites of phytate accumulation in monocotyledonous seeds, whereas globoid crystals of protein bodies are considered to be the site

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

Primary Audience: Broiler Production Managers, Nutritionists, Researchers

MANANGI AND COON: BROILERS AND SOYBEAN MEAL SOURCES

individual enzyme depends on its biochemical properties (pH optimum, enzyme specificity, etc). The relative activity of 6-phytase (from wheat) has a pH optimum of 5, whereas 3-phytase (Aspergillus niger) has pH optima of 2.5 and 5.5 [12]. Certain phytase enzymes are active over a wide pH range (2 to 5.5). The acid phosphatase isolated [13] from Eschericha coli has 8-fold higher specific activity for phytate compared with that of Aspergillus niger phytase [14]. Phytase from E. coli has also been shown to have high proteolytic stability compared with other phytase (from Aspergillus and bacillus) preparations [15]. A comparative study [16] of E. coli phytase and Aspergillus niger phytase in broilers, layers, and pigs indicated a significant (P < 0.05) increase in the availability of phytate P. The prececal digestibility of phytate P due to E. coli phytase was shown to increase by 18% in broilers and 7% in layers compared with P digestibility from Aspergillus phytase. Soybean meal as a major protein source in corn-soybean meal diets for broilers contains 50 to 60% of the total P in the form of phytate P [4, 17]. The P equivalency values for the different microbial phytase enzymes added to poultry feed [18, 19, 20, 21] help determine the amount of NPP that can be replaced in the poultry diet. The ability of microbial phytase to increase P retention in poultry has been tested by incorporating different levels of phytase enzyme, Ca, NPP, total P, and different sources of feed phosphates in poultry diets [21, 22, 23, 24, 25] along with total P or phytate P disappearance in individual feed ingredients, such as corn and soybean meal [4]. Because soybean meal contains a high percentage of phytate P and is a major component of poultry diets, the present study was undertaken to determine 1) the range of total P retention and phytate P disappearance from soybean meal samples collected from different soy crushing plants across the United States and 2) if there is a correlation between various components of soybean meal samples and percentage of total P retention and phytate P disappearance with and without added dietary microbial phytase (Phyzyme XP 5000G). The increased phytate P disappearance values for experimental soybean diets with added phytase will indicate how much phytate P can be made available to poultry through P equivalency value of the enzyme.

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

of phytate storage in dicotyledonous seeds [1], including oil seeds [2]. About 60 to 70% of P in plant feedstuffs is in the form of phytate P, and phytate P content of dry beans depends on the climatic conditions, soil type, and soybean variety. The phytate P content of soybean meal has been reported to range from 0.37 to 0.42% [3]. Dietary phytase has been shown to increase phytate P hydrolysis up to 72.4% (35% with no enzyme) with total P retention of 58% (27% with no enzyme) for broilers fed diets with soybean meal [0.5% Ca and 0.103% nonphytate P (NPP)] as the only source of phytate P [4]. About 6% improvement in total P retention due to dietary phytase supplementation has been reported by Yi et al. [5] using a semipurified diet with soybean meal as the only source of phytate P. The extent of P retention varies depending on the amount of Ca, total P, and phytate P in the diet. Van Der Klis and Versteegh [6] reported 69% phytate P degradation from a soybean meal diet containing 0.5% Ca and 0.18% available P along with a dietary phytase enzyme. Nonruminant animals, such as pigs and poultry, lack significant amounts of endogenous phytase that will hydrolyze phytic acid [7, 8]. To supply the P requirement of an animal, a NPP source needs to be added to the feed or a combination of NPP and a commercial phytase for phytate P hydrolysis. Feeding only a NPP source to poultry to provide 100% of the available P requirement may contribute to environmental pollution because of high levels of P in poultry waste. The ability to utilize P from phytate P helps retain more of the dietary P in reducing dietary inorganic P supplementation and environmental pollution. Phytate can also form complexes with proteins and starch besides forming complexes [1] with metal ions such as Ca, magnesium, iron, and zinc, thereby pronouncing its antinutritional effect. Hydrolysis of phytate reduces the antinutrient effect by releasing phytate bound protein and a variety of metal ions. Phytate P from plant sources can efficiently be made available to simple-stomach animals by using microbial phytase [9, 10, 11]. The enzyme phytase (myo-inositol hexakisphosphosphate 3and 6-phosphohydrolase) from yeasts, fungi, and bacteria hydrolyzes phytate P into inorganic P and inositol phosphate. Many microorganisms have been screened in isolating phytases for use in the poultry and swine industry. The efficacy of an

293

JAPR: Research Report

294 Table 1. Experimental diets1 2

Ingredient

30.000 5.327 2.212 0.204 43.351 8.183 0.002 6.704 2.002 1.050 0.305 0.660 100.000 0.506 0.126 0.192 0.582 <0.010

1

Analyzed values of all 18 experimental diets for total P (%), phytate P (%), Ca (%), phytase (U/kg of diet), crude protein (%), energy (kcal/kg of diet), and DM (%) ranged from 0.19 to 0.23, 0.10 to 0.13, 0.48 to 0.54, 647 to 1,090, 20.2 to 22.9, 4,110 to 4,171, and 90.61 to 92.40, respectively. 2 All ingredients except soybean meal were procured in the form of a basal mix (chick) from Harlan Teklad [27]. The basal mix was formulated to add with a standard NRC solvent-extracted soybean meal to provide the metabolizable energy, protein, amino acids, vitamins, and minerals to supply more than the minimum suggested NRC [30] requirements for 21-d-old broilers. 3 Vitamin mix provided per kg of diet: vitamin A (vitamin palmitate), 10,150 IU; vitamin D3, 3,045 ICU; vitamin E acetate, 50.79 IU; niacin, 60.9 mg; calcium pantothenate, 30.38 mg; riboflavin, 15.26 mg; pyridoxine (pyridoxine HCl), 15.26 mg; thiamine HCl, 20.44 mg; menadione (menadione sodium bisulfite complex), 16.24 mg; folic acid, 4.06 mg; biotin, 3.08 mg; vitamin B12 (0.1% in mannitol), 51.1 mg; sodium selenite, 0.56 mg.Trace mineral mix provided per kilogram of diet: manganese (MnSO4ⴢH2O), 315 mg; zinc carbonate, 203 mg; iron (ferric citrate), 714 mg; copper (CuSO4ⴢ5H2O), 71.4 mg; iodine (KIO3), 4.06 mg; magnesium (MgO), 840 mg.

MATERIALS AND METHODS Male broiler chicks (Cobb 500) [26] were offered a standard starter diet in floor pens until 21 d of age. Chicks of uniform body weight were then placed in individual metabolic cages and offered the test diets (Table 1) consisting of fixed levels of total P combined with a constant level (0.5%) of Ca (Table 1). Although the commercial poultry industry uses 0.9% Ca, in the present study the Ca level of 0.5% was intentionally maintained in all the test diets to maximize total P

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

Test soybean meal Egg white solids, spray-dried Gelatin DL-Methionine Dextrose, monohydrate Corn oil Ethoxyquin (antioxidant) Cellulose Celite Calcium carbonate Sodium chloride Vitamins and minerals3 Total Calculated nutrients Ca Phytate P Total P Analyzed nutrients for basal mix2 (chick) Ca Total P

%

retention and phytate P disappearance. Eighteen of 25 soybean meal samples (Table 2) collected from active soybean crushing plants were selected, analyzed, and mixed with a standard synthetic premix [27]. The 18 soybean meal samples were selected to provide the largest range of P values for appropriate crude protein specified for high-protein dehulled soybean meal. Protein solubility of soybean meal samples was determined by the procedure of Araba and Dale [28]. The urease activity in the soybean meal samples was determined according to the American Association Cereal Chemists method 22-90 [29]. The premix complemented all test soybean meals to provide a balanced diet containing a minimum of required amino acids and essential nutrients (even for the lowest protein soybean meal) as suggested by NRC [30]. The 36 diets consisted of 30% of each test soybean meal and 70% synthetic premix containing a Celite [31] marker (Table 1). A microbial phytase (Phyzyme XP 5000G) [32] was added at 1,000 phytase units (FTU)/kg to one-half of all experimental diets containing each of the test soybean meal samples. Each test soybean meal was also added to the dietary basal without added phytase. The first 8 selected soybean meal samples were used in Experiment 1, and the next 10 of 18 selected soybean meal samples were used in Experiment 2. Ten broiler chicks were fed each of the 36 test diets, thus providing 160 and 200 experimental units in Experiments 1 and 2, respectively. Chicks were acclimated to the cages and diets for 3 d prior to a 2-d excreta collection period. A sample of excreta was collected from each of the 2 collection days, which made a total of 720 excreta samples from both experiments. The excreta from individual birds were collected on trays, frozen (−20°C), and freeze-dried for further analysis. Birds had unlimited access to feed and water for the entire duration of Experiments 1 and 2. Feed consumption and weight gain were recorded during the entire 5-d period of the experiments. Animal use protocol No. 03008 for experiments was approved by the University of Arkansas Institutional Animal Care and Use Committee (IACUC). Diets and excreta were analyzed for total P and Ca by inductively coupled plasma emission spectroscopic method as mentioned by Leske and Coon [25]. Acid-insoluble ash was determined in experimental diets and excreta samples using dry

4.20 4.06 4.05 4.04 4.01 4.01 3.98 3.93 3.44 3.52 3.71 3.73 3.75 3.78 3.78 3.84 3.87 3.92 3.85 3.24 4.19 4.12 3.47 4.08 3.44

0.16 0.20 0.13 0.24 0.12 0.23 0.14 0.18 0.02 0.19 0.04 0.03 0.16 0.08 0.16 0.13 0.02 0.09 0.08 0.17 0.11 0.08 0.07 0.06 0.08

7.39 7.32 6.98 6.46 8.55 8.72 7.32 6.36 5.91 5.89 7.21 7.02 7.61 6.45 6.14 6.92 6.13 6.68 7.11 7.65 7.70 7.07 6.03 7.34 7.20

Total P, mg/g

All values are expressed as an air-dry basis.

1

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25

Source

Phytate P SD 0.35 0.39 0.49 0.07 0.70 0.61 0.40 0.27 0.15 0.08 0.84 0.29 0.61 0.31 0.16 0.84 0.08 0.06 0.66 0.71 0.95 0.06 0.34 0.79 0.39

P SD 3.52 3.91 3.06 3.87 4.02 4.34 4.11 3.58 3.11 3.04 3.24 4.01 3.21 2.93 4.55 3.29 4.17 5.42 4.31 4.21 3.68 3.61 2.83 3.41 4.70

Ca, mg/g 0.12 0.19 0.18 0.14 0.32 0.29 0.28 0.05 0.03 0.03 0.40 0.16 0.22 0.06 0.40 0.42 0.35 0.20 0.39 0.38 0.43 0.07 0.05 0.28 0.46

Ca SD 86.10 87.14 86.81 81.52 88.68 89.33 84.86 84.65 86.11 84.21 84.96 85.95 83.87 87.55 80.28 85.74 86.06 86.87 86.81 85.62 86.96 84.98 84.50 84.95 84.72

DM, % 3.06 2.15 0.69 1.66 0.47 0.78 2.76 2.83 2.05 2.50 1.70 1.72 1.51 1.12 1.36 1.01 1.13 0.89 3.00 1.40 0.61 2.67 3.25 1.06 1.59

DM SD 48.85 48.57 49.15 47.88 51.69 51.57 47.71 49.34 49.21 49.04 47.66 50.91 48.67 49.30 47.89 48.22 48.07 50.32 44.34 46.59 42.85 49.00 47.52 49.59 40.44

Protein, % 0.48 0.27 0.26 0.32 0.22 0.39 1.02 0.18 0.22 0.21 0.36 0.55 0.21 0.38 0.37 0.30 0.36 0.26 0.62 0.57 0.51 0.21 0.20 0.42 0.46

Protein SD 10.17 8.68 9.34 11.11 8.39 9.03 7.98 7.78 9.68 8.32 10.94 8.35 10.06 10.48 10.57 7.92 9.52 13.04 16.09 9.48 11.36 8.64 7.90 9.98 10.38

NDF, %

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

Phytate P, mg/g

Table 2. Analysis of soybean meal samples collected from various sources within the United States1 Urease index, U of pH increase 0.04 0.03 0.03 0.07 0.04 0.04 0.06 0.02 0.07 0.04 0.03 0.02 0.05 0.04 0.05 0.21 0.03 0.06 0.22 0.09 0.05 0.02 0.01 0.25 0.05

NDF SD 0.84 0.51 0.69 0.56 0.74 0.64 0.52 0.56 0.74 0.29 0.68 0.58 0.41 0.66 1.41 0.52 0.66 4.08 0.96 0.39 0.27 0.46 0.66 0.95 0.92

72.50 79.80 77.40 80.70 80.40 77.10 86.40 70.00 71.40 71.10 74.10 78.70 71.90 73.10 68.80 82.80 72.30 65.10 83.71 85.29 79.86 69.98 76.36 79.43 68.46

Protein solubility, %

MANANGI AND COON: BROILERS AND SOYBEAN MEAL SOURCES 295

JAPR: Research Report

296

Statistical Analysis Data were subjected to 2-way ANOVA and t-test to determine statistical significance [37]. Data from trial 1 and 2 were pooled for t-test because the preliminary statistical analysis of data from both the trials indicated no significant (P > 0.05) interaction of trial × enzyme.

RESULTS AND DISCUSSION The present bioassay was conducted in broilers to evaluate the effect of different sources of soybean meal on performance and retention of various nutrients in general and phytate P disappearance in particular using microbial phytase. The analysis of soybean meal samples collected from 25 active soybean crushing plants in the United States indicated (Table 2) the phytate P, total P, Ca, DM, protein, and NDF contents ranged from 0.32 to 0.42, 0.59 to 0.87, 0.28 to 0.54, 80.28 to 89.33, 40.44 to 51.69, and 7.78 to

16.09%, respectively. The urease activity values of various soybean meal samples (Table 2) used in Experiments 1 and 2 ranged from 0.02 to 0.21. The protein solubility (in 0.2% potassium hydroxide) of various soybean meal sources used in Experiments 1 and 2 (Table 2) ranged from 65.1 to 86.4%. The effects of dietary phytase on chick performance, P intake and excretion during 5-d chick bioassays are presented in Table 3 and 4 for Experiments 1 and 2, respectively. The main effect of phytase (1,000 U/kg of diet) indicates a significant (P < 0.05) increase in body weight gain and improvement in feed to gain ratio during 5 d bioassays (Tables 3 and 4). Feed intake was not affected (P > 0.05) by phytase supplementation (Tables 3 and 4). Body weight, feed consumption, and feed:gain ratio were not influenced (P > 0.05) by soybean meal sources. The interactions between soybean meal sources and phytase were also nonsignificant (P > 0.05). Pooled mean comparisons for 18 test soybean meal sources for phytase added groups compared with groups fed diets without phytase (Table 5) indicate a significant 23-g improvement in body weight (P < 0.05) and a 16-point improvement in feed conversion (P < 0.0001). The pooled mean comparison of feed consumption for broilers fed the different soybean meal sources with and without phytase was not significantly different (P > 0.05). The response on broiler performance (body weight, feed consumption, and FCR) with dietary phytase supplementation in the study reported herein is in agreement with the findings of other research on feeding phytase to broilers and turkey poults [22, 38, 3, 18, 39, 40, 41, 42, 43, 24]. The improvement in body weights and performance in broilers caused by the addition of phytase in feed could be attributed to 1) greater availability of P, especially myo-inositol, released by phytate hydrolysis or dephosphorylation; 2) release of trace elements or minerals from complexes with phytate; 3) possible increase in starch digestibility [44]; and 4) increased availability of amino acids or protein from protein-phytate complexes that occur naturally in feed ingredients [45]. The 5-d bioassay used in the present work was previously developed by Leske and Coon [4], and the assay was mainly used to study retainable P from nonphytate sources.

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

ash and hydrochloric acid digestion technique of Scott and Balnave [33]. Diet and excreta phytate P were measured as inositol hexaphosphate by using ion-exchange chromatography as described by Bos et al. [34]. Feed and excreta N and moisture were determined by standard AOAC procedures [35]. Neutral detergent fiber (NDF) of soybean meal samples was quantified by a semiautomated fiber extraction system using a filter bag technique [36]. No sulfite or heat-stable amylase was included in the procedures to determine NDF of soybean meal. The gross energy of test feed and gross energy of excreta were measured with a Parr bomb calorimeter. Phytate P disappearance and retention values of total P, NPP, N, and energy were determined using the diet and excreta phytate P, total P, N and gross energy, and acid-insoluble ash concentrations with a marker digestibility equation reported by Scott and Balnave [33]. The term disappearance is used instead of retention for phytate P because there is no established way to confirm that all of the retained P from phytate is from phytate P hydrolysis. The total excreta P that was measured by inductively coupled plasma emission could have been a combination of P from phytate P hydrolysis and from traditional NPP source. Experimental diets were assayed for phytase from Danisco Animal Nutrition (Marlborough, UK).

1,000 0

1,000 1,000 1,000 1,000 1,000 1,000 1,000 1,000 0 0 0 0 0 0 0 0 PSEM1

0.5356 0.5524 0.5614

340 333

199a 170b

0.5654 0.0145 0.9400

340 350 346 332 353 320 297 348

340 360 339 360 347 318 334 330 340 338 354 314 360 324 261 369 27

FI, g

191 200 185 171 201 178 156 197

210 211 189 193 211 189 189 199 171 187 180 157 188 162 123 194 23

BWG, g

0.8685 0.0005 0.7642

1.75b 1.99a

1.85 1.78 1.98 1.96 1.84 1.84 1.86 1.84

1.66 1.74 1.92 1.87 1.68 1.69 1.83 1.68 2.04 1.83 2.05 2.02 2.05 2.04 1.90 2.04 0.13

FCR

0.8636 0.5191 0.5486

752 734

745 786 760 724 751 757 683 724

744.5 807.3 744.7 784.6 738.8 751.1 767.1 686.0 744.9 759.9 778.2 684.0 767.0 765.3 598.7 769.0 60

Total P intake, mg

P 0.6166 0.5328 0.5762

419 410

434 420 426 422 432 413 366 400

433.3 431.9 417.8 457.7 424.3 409.3 411.3 379.0 433.6 406.5 436.6 399.0 440.5 417.0 321.0 424.8 34

PP intake, mg

0.3520 0.5029 0.5129

333 323

311 365 334 302 320 345 317 324

311.2 375.4 326.9 327.0 314.5 341.8 355.8 307.1 311.4 353.4 341.6 285.0 326.5 348.3 277.7 344.3 26

NPP intake, mg

Excreta PP DM, mg/g 0.86 0.51 0.77 1.06 0.57 0.61 0.78 0.97 3.11 3.74 3.32 4.15 3.47 3.33 3.12 2.65 0.32 1.99 1.98 1.90 2.91 1.86 1.75 1.95 1.73 0.75b 3.39a 0.3769 <0.0001 0.2575

Excreta P DM, mg/g 3.22 2.74 3.18 3.18 2.75 2.63 3.02 2.98 4.96 5.44 5.18 5.89 4.98 4.70 5.37 4.53 0.27 4.09b 3.97bc 4.07b 4.81a 3.74bc 3.50c 4.20b 3.68bc 2.94b 5.15a 0.0456 <0.0001 0.2871

1

a–c

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

0.6065 0.0035 0.9194

2.19a 1.76b

2.11 1.99 2.17 1.89 1.89 1.75 2.25 1.95

2.36 2.22 2.42 2.13 2.18 2.02 2.24 2.01 1.85 1.71 1.86 1.74 1.51 1.37 2.26 1.88 0.28

Excreta NPP DM, mg/g

Means within a column with differing superscripts are significantly different (P < 0.05). FI = feed intake; BWG = body weight gain; FCR = feed conversion ratio (feed to gain ratio); PSEM = pooled standard error of the mean, PP = phytate P; NPP = nonphytate P.

Source of variation Source Enzyme Source × enzyme

Source effect 1 2 3 4 5 6 7 8 Enzyme effect

1 2 3 4 5 6 7 8 1 2 3 4 5 6 7 8

Source

Phytase, U/kg

Table 3. The effect of dietary phytase on chick performance, P intake and P excretion during a 5 d chick bioassay, Experiment 11

MANANGI AND COON: BROILERS AND SOYBEAN MEAL SOURCES 297

1 2 3 4 5 6 7 8 9 10 1 2 3 4 5 6 7 8 9 10 PSEM1 Source effect 1 2 3 4 5 6 7 8 9 10

Source

1,000 1,000 1,000 1,000 1,000 1,000 1,000 1,000 1,000 1,000 0 0 0 0 0 0 0 0 0 0

403 392 393 369 414 391 422 413 422 409 430 409 396 413 410 385 424 311 429 350 20 415 400 394 393 412 375 423 368 426 378

252 240 247 233 248 221 257 223 249 232

FI, g

253 248 255 228 258 240 263 255 257 256 251 233 234 237 242 203 249 184 242 211 15

BWG, g

1.66 1.70 1.61 1.70 1.67 1.70 1.66 1.68 1.72 1.64

1.62 1.62 1.55 1.62 1.61 1.62 1.62 1.65 1.64 1.60 1.73 1.78 1.70 1.76 1.71 1.77 1.71 1.73 1.78 1.67 0.06

FCR

800 782 851 809 916 773 832 765 859 820

777.6 765.5 848.1 759.5 921.7 806.5 830.3 859.0 850.6 889.0 829.7 799.0 855.7 850.1 912.5 738.4 833.8 647.9 865.0 760.5 42

Total P intake, mg 372.0 367.5 442.3 342.2 417.4 394.6 398.7 400.4 403.3 427.1 396.9 383.6 446.2 383.0 413.3 361.3 400.4 302.0 410.1 365.4 20 383bcd 376bcd 444a 364d 415ab 378bcd 400abcd 357d 407abc 394bcd

417bc 407bc 407bc 445b 501a 395c 432bc 409bc 451b 426bc

NPP intake, mg

405.6 397.9 405.9 417.3 504.3 411.9 431.5 458.6 447.3 461.9 432.8 415.3 409.5 467.1 499.2 377.2 433.3 345.9 454.9 395.1 22

PP intake, mg

2.10 2.35 2.01 1.84 1.95 2.35 1.97 2.03 2.39 2.38 Contined

1.65 1.31 1.55 2.08 2.08 1.49 1.16 1.85 1.52 1.88

3.75b 3.66b 3.56bc 3.92ab 4.02ab 3.84ab 3.13c 3.88ab 3.91ab 4.26a

2.17 2.45 2.24 1.97 2.33 2.30 2.42 2.19 2.57 2.55 2.00 2.24 1.63 1.73 1.70 2.40 1.43 1.83 2.24 2.23 0.20

0.78 0.42 0.78 0.64 0.35 0.47 0.21 0.70 0.84 0.61 2.83 2.21 2.79 3.27 3.16 2.52 2.29 3.29 2.09 3.00 0.23

2.95gh 2.87gh 3.01gh 2.61h 2.68h 2.76gh 2.63h 2.88gh 3.41fg 3.16fgh 4.83abcd 4.45bcd 4.42cd 5.00abc 4.86abcd 4.91abcd 3.73ef 5.12ab 4.33de 5.22a 0.21

Excreta NPP DM, mg/g

Excreta PP DM, mg/g

Excreta P DM, mg/g

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

Phytase, U/kg

Table 4. The effect of dietary phytase on chick performance, P intake, and P excretion during a 5-d chick bioassay, Experiment 21

298

JAPR: Research Report

0.0546 0.3185 0.1071 0.9703 0.0005 0.9994 0.1106 0.3452 0.1030 0.3801 0.0027 0.5592

Means within a column with differing superscripts are significantly different (P < 0.05). FI = feed intake; BWG = body weight gain; FCR = feed conversion ratio (feed to gain ratio); PSEM = pooled standard error of the mean; PP = phytate P; NPP = nonphytate P. 1

Source Enzyme Source × enzyme

403 395 252a 230b 1,000 0 Enzyme effect

a–h

0.0651 0.0004 0.5746 0.0706 <0.0001 0.1823 0.0135 <0.0001 0.0285

P

0.0020 0.3215 0.0964

Source of variation

398 388 432 428 830 817 1.61b 1.73a

FCR Source

0.0071 0.3160 0.1209

0.59b 2.76a 2.91b 4.71a

2.32a 1.96b

Excreta PP DM, mg/g Excreta P DM, mg/g NPP intake, mg PP intake, mg Total P intake, mg FI, g BWG, g Phytase, U/kg

299

In Experiment 1, there was no effect (P > 0.05) of soybean source, enzyme, or source by enzyme on intake of total P, phytate P, or NPP (Table 3). In Experiment 2 (Table 4), soybean meal source did not significantly (P > 0.05) affect total P intake, but intake of phytate P and NPP was significantly (P < 0.05) affected. There was no effect (P > 0.05) of enzyme or interaction of source by enzyme on intake of total P, phytate P, or NPP. Pooled mean comparisons for 18 test soybean meal sources for phytase added groups compared with groups fed diets without phytase (Table 5) indicated a nonsignificant (P > 0.05) effect of phytase on intake of total P, phytate P, and NPP. In both experiments, soybean meal source influenced excreta P significantly (P < 0.05) but not excreta phytate P or NPP. Dietary phytase supplementation significantly (P < 0.05) reduced total P and phytate P and increased NPP in the excreta. The interaction of source by enzyme did not have any influence on excreta total P, phytate P, or NPP. Pooled mean comparisons for 18 test soybean meal sources for phytase added groups compared with groups fed diets without phytase (Table 5) indicated a significant (P < 0.0001) reduction of 1.96 mg of total P and 2.34 mg of phytate P/g of DM excreta and a significant (P < 0.0001) increase of 0.38 mg of NPP/g of DM excreta. In Experiment 1, dietary phytase supplementation significantly (P < 0.0001) increased the percentages of total P retention and phytate P disappearance and reduced (P = 0.0453) the retention percentage of NPP (Table 6). Soybean meal source affected (P < 0.05) total P retention but not phyate P disappearance (P > 0.05) or NPP retention (P > 0.05; Table 6). There was no interaction effect of source by enzyme on total P retention, NPP retention, and phytate P disappearance (Table 6). In experiment 2, phytase significantly (P < 0.0001) increased the percentages of total P retention and phytate P disappearance and nonsignificantly reduced (P = 0.1818) the retention percentage of NPP (Table 7). Soybean meal source did not affect (P > 0.05) total P retention, phytate P disappearance, or NPP retention (Table 7). Interaction between soybean meal source and enzyme had a significant (P < 0.05) effect on total P retention but not on phytate P disappearance and NPP retention (Table 7). Pooled mean compari-

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

Table 4 (Continued). The effect of dietary phytase on chick performance, P intake, and P excretion during a 5-d chick bioassay, Experiment 21

Excreta NPP DM, mg/g

MANANGI AND COON: BROILERS AND SOYBEAN MEAL SOURCES

JAPR: Research Report

300

Table 5. Means comparison between phytase enzyme supplemented and nonsupplemented groups fed diets containing 18 selected soybean meal samples, Experiments 1 and 2 With enzyme, mean1 ± SE 230.00 377.00 1.67 798.00 426.00 371.00 2.92 0.66 2.26 60.24 82.50 32.86 69.59 59.45 11.42

± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

4.98 6.18 0.02 12.48 6.54 6.60 0.05 0.04 0.05 0.79 0.96 1.72 0.57 0.64 0.93

P > |t|

± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

0.0013 0.5136 <0.0001 0.4650 0.6059 0.3766 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 0.0252 0.1381 0.0004 <0.0001

207.00 370.00 1.83 784.00 421.00 363.00 4.88 3.00 1.88 29.44 19.81 38.12 68.15 55.84 −10.36

5.29 7.03 0.03 13.68 7.43 6.85 0.07 0.10 0.06 1.14 1.94 2.39 0.65 0.78 1.05

1

Mean values of upper and lower confidence limits.

sons for 18 test soybean meal sources for phytase added groups compared with groups fed diets without phytase (Table 5) indicated a significant (P < 0.0001) 62.69 percentage points increase in phytate P disappearance and 30.8 percentage points increase in total P retention and a significant (P < 0.05) decrease in NPP retention by 5.26 percentage points. The phytate P disappearance values (pooled means) of this study agree with other previous research reports. Leske and Coon [4] reported 34.9% phytate P hydrolysis in broiler chicks (5d bioassay) fed a semipurified diet containing 0.273% total P, 0.141% phytate P, and 0.4% Ca with no enzyme and 72.4% phytate hydrolysis with 600 FTU phytase/kg. Shirley and Edwards [24] also reported phytate P disappearance of 65.2% in the excreta with 1,500 U of phytase/kg of diet compared with 40.3% phytate P disappearance with no dietary phytase supplementation in broilers fed corn-soybean meal basal diets containing 0.88% Ca, 0.46% total P, and 0.27% phytate P. Simons et al. [22] reported 62.5% total P availability with supplementation of 1,000 U of phytase/kg of diet compared with 49.8% in the control group with no phytase supplementation in broilers fed corn-sorghum-soy diets with 0.6% Ca, 0.45% total P, and 0.3% phytate P. Qian et al. [46] reported a linear increase in P retention from 52.9 (control) to 58.4% with phytase supplementation. Broz et al. [47] reported

a significant improvement in apparent availability of P from 44.32 (control) to 52.71% with dietary phytase supplementation. Viveros et al. [43] reported a significant increase in P retention from 51.47 (control) to 57.68% with phytase. The total P retention shown in Table 5 was in close agreement with the study by Leske and Coon [4] that reported 27 and 58% total P retention with and without phytase, respectively, in chicks fed semipurified diets. The variation in the level of phytate hydrolysis and total P retention may be due to the percentage of inclusion levels of total P, phytate P, and Ca and the type of feed ingredients (corn, soybean meal, wheat midds, canola meal, barley, and sorghum), age of birds, and the duration of trial. In the present study the total P and phytate P levels were maintained at low levels purposely so that chicks could optimize the use of released P from phytate hydrolysis, increase phytase efficiency in hydrolyzing phytate P, and increase the maximum percentage of phytate P hydrolysis in broilers fed diets with added phytase and different soybean meal sources. The retention of NPP for all groups fed phytase was significantly lower than observed for NPP retention for chick groups that were not fed phytase. The lower retention of NPP for these phytase fed groups is probably because the optimum retention of NPP is higher for chicks fed low dietary P levels. The chicks fed phytase diets received an average of 0.075% additional retainable P above the feed

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

Body weight, g Feed consumption, g Feed conversion ratio P intake, mg Phytate P intake, mg Nonphytate P intake, mg Excreta P, mg/g of DM Excreta Phytate P, mg/g of DM Excreta nonphytate P, mg/g of DM Total P retention, % Phytate P disappearance, % Nonphytate P retention, % Gross energy retention, % N retention, % Ca retention, %

Without enzyme, mean1 ± SE

MANANGI AND COON: BROILERS AND SOYBEAN MEAL SOURCES

301

Table 6. The effect of dietary phytase in a 5-d chick bioassay on the retention and disappearance of P, gross energy, N, and Ca, Experiment 11

Source

1,000 1,000 1,000 1,000 1,000 1,000 1,000 1,000 0 0 0 0 0 0 0 0

1,000 0

Total P retention, %

PP disappearance, %

GE retention, %

N retention, %

Ca retention, % 17.76 16.79 6.45 14.52 8.54 17.54 21.79 16.19 −3.11 −6.95 −4.81 −1.19 −10.87 2.17 −9.87 −5.43 3.74

63.00 66.47 56.81 57.14 61.08 68.62 63.73 60.52 31.66 32.51 31.60 28.78 29.57 48.41 25.88 32.29 3.84

83.25 88.26 81.60 76.18 86.25 87.12 83.32 77.31 28.00 13.64 23.90 14.14 15.80 32.81 21.67 27.89 6.36

34.81 41.42 25.13 30.48 27.13 46.48 41.10 39.79 36.76 54.22 41.43 49.28 48.15 67.09 30.75 37.72 9.55

76.01a 73.35abc 69.39bcd 70.36abcd 70.09abcd 70.69abcd 73.09abcd 72.61abcd 68.45cd 73.52abc 70.32abcd 76.15a 71.09abcd 75.49ab 66.60d 68.20cd 2.29

63.87 61.81 58.71 54.38 55.10 64.00 63.00 58.90 54.39 60.74 57.12 58.80 54.04 63.82 54.14 52.29 2.76

47.33bc 51.04b 45.60bc 40.13c 47.08bc 60.20a 44.81bc 47.69bc

55.63 54.34 55.95 38.96 54.94 64.49 52.50 54.85

35.79 47.24 32.38 41.76 36.47 55.07 35.92 38.85

72.23 73.42 69.80 73.84 70.54 72.69 69.84 70.61

59.13abc 61.32ab 58.00bc 57.03bc 54.63c 63.93a 58.63abc 55.89bc

7.33 6.00 1.45 5.09 −0.09 11.14 5.96 6.36

62.30a 32.77b

83.26a 22.16b

36.53b 46.21a

71.95 71.81

60.25a 57.11b

14.99a −4.63b

0.0024 <0.0001 0.4457

0.4954 <0.0001 0.4952

Source of variation Source Enzyme Source × enzyme

NPP retention, %

P 0.2075 0.0453 0.6599

0.4964 0.5353 0.0352

0.0198 0.0324 0.1974

0.1142 <0.0001 0.3074

Means within a column with differing superscripts are significantly different at P < 0.05. PP = phytate P; NPP = nonphytate P; GE = gross energy; PSEM = pooled standard error of the mean.

a–d 1

NPP based on phytate hydrolysis and disappearance, which might have provided enough additional P to pass the physiological threshold. The broilers that received P levels that were higher (through phytate P hydrolysis) than the physiological threshold needed for maximum use and retention were eliminating the additional P most likely through the kidney [25]. The percentage of NPP in the excreta of chicks fed phytase was always higher than the comparable P group fed diets without phytase. The higher percentages of NPP levels in the excreta for the phytase fed broilers might also have been due to no direct measurement of NPP. The percentage of NPP was mea-

sured by the difference between total P and phytate P with the assumption of phytate P being an IP6 structure. In Experiment 1, soybean meal source did not have a significant (P > 0.05) effect on gross energy retention but did influence (P < 0.05) N retention (Table 6). Phytase supplementation significantly increased N retention (P < 0.05) by 3.14 percentage points but had no effect on gross energy retention (P > 0.05). Interaction of source by enzyme significantly (P < 0.05) influenced gross energy retention but not retention of N. In experiment 2, soybean meal source did not have a significant (P > 0.05) effect on N retention but

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

1 2 3 4 5 6 7 8 1 2 3 4 5 6 7 8 PSEM Source effect 1 2 3 4 5 6 7 8 Enzyme effect

Phytase, U/kg

JAPR: Research Report

302

Table 7. The effect of dietary phytase in a 5-d chick bioassay on the retention and disappearance of P, gross energy (GE), N, and Ca, Experiment 21

Source

1,000 1,000 1,000 1,000 1,000 1,000 1,000 1,000 1,000 1,000 0 0 0 0 0 0 0 0 0 0

1,000 0

Total P retention, %

PP disappearance, %

NPP retention, %

GE retention, %

Ca retention, %

60.01a 58.65a 62.79a 64.92a 57.83a 63.29a 61.48a 62.67a 59.93a 61.02a 26.86de 32.58cd 40.22bc 21.35e 35.53bcd 30.01cde 44.54b 32.90cd 25.19de 25.59de 3.42

80.15 88.88 80.09 84.52 91.31 87.86 94.08 83.68 78.71 85.92 19.21 37.13 21.88 15.56 24.69 31.30 34.78 19.24 33.48 20.15 5.53

38.04 25.92 46.90 41.10 32.44 37.64 26.20 38.62 36.56 34.10 35.20 27.70 57.04 39.32 48.62 28.66 55.11 48.54 18.53 46.56 6.91

73.70 68.53 72.92 69.68 61.06 71.51 67.20 71.43 68.24 70.68 70.00 68.40 69.96 65.74 69.96 70.28 68.88 73.23 60.52 66.53 2.11

61.70 60.27 60.19 55.83 50.29 58.07 57.63 59.97 61.35 60.42 58.00 58.15 57.94 49.34 58.00 52.35 57.08 57.26 49.15 53.52 2.57

4.51 6.62 11.24 14.61 8.03 9.37 14.22 7.85 8.13 3.54 −12.84 −8.15 −9.95 −14.87 −16.41 −18.26 −8.93 −17.40 −16.52 −19.76 3.30

45.80 45.61 54.10 41.15 44.11 46.65 53.78 49.44 39.09 42.12

54.04 63.01 57.71 42.02 50.31 59.58 67.12 55.04 54.04 50.85

36.82 26.79 50.80 40.09 43.23 33.15 39.34 43.03 26.54 39.85

72.11a 68.46abc 71.78a 67.53abc 66.54bc 70.90ab 67.97abc 72.23a 64.03c 68.46abc

60.12 59.21 59.32 52.29 55.03 55.21 57.38 58.77 54.70 56.74

−2.93 −0.76 3.09 −0.13 −7.01 −4.44 3.70 −3.37 −5.56 −9.17

61.25a 31.02b

85.19a 24.65b

36.09 40.58

69.86 68.11

58.88a 55.02b

8.78a −14.33b

0.0981 <0.0001 0.0451

0.0731 <0.0001 0.4759

Source of variation Source Enzyme Source × enzyme

N retention, %

P 0.0727 0.1818 0.2145

0.0154 0.2936 0.0682

0.2032 0.0095 0.1334

0.1683 <0.0001 0.6849

Means within the column with differing superscripts are significantly different at P < 0.05. PP = phytate P; NPP = nonphytate P; PSEM = pooled standard error of the mean.

a–e 1

did affect (P < 0.05) gross energy retention (Table 7). Phytase supplementation significantly increased N retention (P < 0.05) by 3.86 percentage points but had no effect on gross energy retention (P > 0.05). The interaction of source by enzyme did not have a significant (P > 0.05) effect on retention of gross energy and N. Pooled mean comparisons for 18 test soybean meal sources for phytase added groups compared with groups fed

diets without phytase (Table 5) indicated a significant (P < 0.001) 3.61 percentage points increase in N retention. The increase of 3.61 percentage points in N retention and 1.44 percentage points in energy retention from both experiments in phytase-supplemented chicks compared with that of unsupplemented chicks could have been due to the ability of phytase to free the complexes and nul-

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

1 2 3 4 5 6 7 8 9 10 1 2 3 4 5 6 7 8 9 10 PSEM Source effect 1 2 3 4 5 6 7 8 9 10 Enzyme effect

Phytase, U/kg

MANANGI AND COON: BROILERS AND SOYBEAN MEAL SOURCES

303

Table 8. The amount of dietary retainable P produced from different soybean meals with phytase supplementation at 1,000 phytase units/kg of diet, Experiments 1 and 2

Diet

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 Average

1 2 3 4 5 6 7 8 1 2 3 4 5 6 7 8 9 10

PP disappearance, gain %

Dietary retainable P, gain %

0.13 0.12 0.12 0.13 0.12 0.13 0.12 0.12 0.10 0.10 0.10 0.11 0.12 0.11 0.10 0.11 0.11 0.11 0.11

55.25 74.62 57.70 62.04 70.45 54.31 61.65 49.42 60.94 51.75 58.21 68.96 66.62 56.56 59.30 64.44 45.23 65.77 60.18

0.078 0.097 0.075 0.087 0.092 0.077 0.080 0.064 0.067 0.057 0.064 0.083 0.088 0.068 0.065 0.078 0.054 0.079 0.075

1

Treatments 1 through 8 are from Experiment 1, and Treatments 9 through 18 are from Experiment 2. Phytate P.

2

lify the negative effect of phytate because phytate can form metallic complexes, protein complexes and complexes with lipids [1] and can inhibit αamylase digestion of starch [44]. The literature indicates that about 4.5% improvement in AME or AMEn (kcal/kg) with 750 to 800 U of dietary phytase supplementation [24, 48] compared with unsupplemented or control groups. Contrarily, there is some evidence [49, 50] that dietary phytase supplementation has no significant effect on energy use. Coon and Manangi [51] reported an

improvement of 1.9 to 6 percentage points in ileal digestibility of total amino acids with the addition of 500 to 1,000 U of phytase/kg of corn-soybean meal diet containing 0.49% total P and 0.24% phytate P. Kies et al. [12] mentioned in their review that the digestibility of amino acids and crude protein improve by 1 to 3% with the addition of microbial phytase. The present study indicates strong evidence for different responses to phytase when chicks are fed different soybean meal samples based on a significant source ×

Table 9. Correlations between the components of soybean meal samples and percentages of total P retention and of phytate P (PP) disappearance by chicks, Experiment 1 With enzyme Total P retention, % PP disappearance, % Without enzyme Total P retention, % PP disappearance, %

Total P r (P r (P

= = = =

0.325 0.0356)* 0.464 0.002)*

r (P r (P

r (P r (P

= = = =

0.355 0.0287)* 0.098 0.5592)

r = −0.033 (P = 0.8444) r = 0.001 (P = 0.9976)

NDF = neutral detergent fiber. *Values are significant (P < 0.05).

1

PP = = = =

0.024 0.8826) 0.143 0.3679)

Ca r (P r (P

= = = =

0.372 0.0152)* 0.288 0.0644)

r = 0.236 (P = 0.1535) r = −0.020 (P = 0.9054)

Protein

NDF1

r (P r (P

= = = =

0.185 0.2409) 0.267 0.0872)

r = −0.137 (P = 0.3883) r = −0.125 (P = 0.4290)

r (P r (P

= = = =

0.402 0.0124)* 0.150 0.3687)

r = −0.038 (P = 0.8228) r = −0.085 (P = 0.6116)

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

Treatment1

Dietary PP,2 %

JAPR: Research Report

304

Table 10. Correlations between the components of soybean meal samples and percentages of total P retention and phytate P (PP) disappearance by chicks, Experiment 2 With enzyme Total P retention, % PP disappearance, % Without enzyme Total P retention, % PP disappearance, %

Total P

PP = = = =

Ca

Protein

NDF1

r = 0.319 (P = 0.0166)* r = −0.119 (P = 0.3764)

r (P r (P

0.143 0.2935) 0.125 0.3535)

r = −0.035 (P = 0.8007) r = 0.149 (P = 0.2679)

r = 0.007 (P = 0.9585) r = −0.002 (P = 0.9882)

r = −0.006 (P = 0.9662) r = 0.014 (P = 0.9179)

r = 0.151 (P = 0.2610) r = −0.218 (P = 0.1033)

r = −0.008 (P = 0.9504) r = −0.075 (P = 0.5806)

r = −0.082 (P = 0.5440) r = −0.143 (P = 0.2895)

r = −0.349 (P = 0.0078)* r = −0.259 (P = 0.0519)

r = 0.041 (P = 0.7643) r = −0.148 (P = 0.2727)

Neutral detergent fiber. *Values are significant (P < 0.05).

enzyme interaction for energy retention (P = 0.0352 in Experiment 1 and P = 0.0682 in Experiment 2) and low probabilities for N retention (P = 0.1974 in Experiment 1 and P = 0.1334 in Experiment 2). Further, the correlation between N and energy retention vs. various soybean meal components indicates a significant (P = 0.0381) correlation (r = 0.3377) between energy retention and percentage of NDF in Experiment 1 and a significant (P = 0.048) correlation (r = 0.2586) between energy retention and trypsin inhibitor in Experiment 2. Dietary phytase supplementation caused significant (P < 0.0001) improvement in Ca retention in both experiments (Tables 6 and 7). Neither source nor interaction of source by enzyme had any effect (P > 0.05) on Ca retention. Pooled mean comparisons for 18 test soybean meal sources for phytase added groups compared with groups fed diets without phytase (Table 5) indicated a significant (P < 0.0001) increase in Ca retention by 21.78 percentage points. The Ca retention values for each of the research experiments from our lab (unpublished) show there is a minimum dietary requirement for retainable P of 0.0875% to maintain a positive Ca balance and a requirement of 0.255 to 0.28% retainable P to optimize Ca retention. The percentages of dietary retainable P gain or P equivalency values for phytase (at 1,000 U/ kg) in Experiments 1 and 2 are presented in Table 8. The values for percentage of dietary retainable P ranged from 0.054 to 0.097. The P equivalency values of the phytase in the present research are in agreement with P equivalent values of microbial phytases pre-

viously reported by other researchers. The microbial phytase can be supplied as a replacement to inorganic P. Supplementation of 600 U of phytase/kg of feed has been shown to replace 0.1% inorganic P in broiler chicks [20]. Schoner et al. [19] showed that 700 U of microbial phytase is equivalent to 1 g of P in the form of monocalcium phosphate based on P retention in broilers fed a corn-soybean meal diet for 2 wk. Kornegay et al. [21] reported that 939 U of microbial phytase is equivalent to 1 g of P from defluorinated phosphate in broilers fed corn-soybean meal diets with graded levels of microbial phyatase and a fixed level of phytate P (0.24%) in combination with 0.2, 0.27, and 0.34% NPP and 0.88, 1.02, and 1.16% Ca, respectively. The small discrepancy or variation in the P equivalency of phytase among various studies (19, 20, 21) in comparison with those in the present work could mainly be attributed to source of microbial phytase and its efficacy in the biological system, source and amount of dietary phytate, amount of dietary Ca and available P, and age of birds or period of study. In the present study, the retainable P was solely from soybean meal because the semipurified diet was used to determine the phytase effect and the extent of phytate hydrolysis caused by addition of phytase. In Experiments 1 and 2, the feed components (total P, phytate P, Ca, protein, and NDF) were tested for correlation (for P < 0.05) with percentages of total P retention and phytate P disappearance in chicks (Tables 9 and 10). In Experiment 1, the significant correlations in chicks fed phytase were found between total P and percentage of total P retention (r = 0.325, P = 0.0356), between

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

1

MANANGI AND COON: BROILERS AND SOYBEAN MEAL SOURCES

between protein and percentage of total P retention (r = 0.349, P = 0.0078). The amount of protein in soybean samples in Experiments 1 and 2 had a consistent correlation to the percentage of total P retention when the chicks were not fed phytase. The correlations between some of the feed components and percentages of total P retention and phytate P disappearance in Experiments 1 and 2 were not the same. The low correlation values for those that were significant in both experiments indicate the significant correlations may be due to chance.

CONCLUSIONS AND APPLICATIONS 1. Chicks fed 18 different sources of soybean meal with added phytase at 1,000 FTU /kg of diet with 0.5% Ca provided an average of 0.07% retainable P equivalents from phytate P hydrolysis and use. 2. The results indicate no correlation between components (total P, phytate P, Ca, protein, and NDF) of soybean meal samples and percentages of phytate P disappearance and total P retention for groups fed diets with or without added phytase. 3. Dietary supplementation of 1,000 FTU/kg of diet increased the phytate P disappearance by 62.69 percentage points and retention of total P, gross energy, N, and Ca by 30.8, 1.44, 3.61, and 21.78 percentage points, respectively. 4. The 5-d bioassay utilized in the present experiment was intentionally designed to maximize total P retention and phytate P disappearance by feeding a 2.5:1 ratio of Ca to total P and a 3.85:1 ratio of Ca to phytate P. 5. Soybean meals may show different responses to phytase with regard to improving N and energy use.

REFERENCES AND NOTES 1. Cosgrove. 1980. Inositol Phosphates: Their chemistry, Biochemistry and Physiology. Elsevier Sci. Publ. Co., New York, NY. 2. Erdman, J. W., Jr. 1979. Oilseed phytates: Nutritional implications. J. Am. Oil Chem. Soc. 56:736–741. 3. Ravindran, V., W. L. Bryden, and E. T. Kornegay. 1995. Phytates: Occurrence, bioavailability and implications in poultry nutrition. Poult. Avian Biol. Rev. 6:125–143. 4. Leske, K. L., and C. N. Coon. 1999. A bioassay to determine the effect of phytase on phytate phosphorus hydrolysis and total phosphorus retention of feed ingredients as determined with broilers and laying hens. Poult. Sci. 78:1151–1157. 5. Yi, Z., E. T. Kornegay, and D. M. Denbow. 1996. Supplemental microbial phytase improves zinc utilization in broilers. Poult. Sci. 75:540–546. 6. Van der Kliss, J. D., and H. A. J. Versteegh. 1996. Phosphorus nutrition of poultry. Pages 71–83 in Proc. Nottingham Feed Manufacturers Conf., Nottingham, UK. 7. Cooper, R. J., and H. S. Gowing. 1983. Mammalian small intestine phytase (EC 3.1.3.8). Br. J. Nutr. 50:429–435. 8. Maenz, D. D., and H. L. Classen. 1998. Phytase activity in the small intestinal brush border membrane of the chicken. Poult. Sci. 77:557–563.

9. Nelson, T. S., T. R. Shieh, R. J. Wodzinski, and J. H. Ware. 1971. Effect of supplemental phytase on the utilization of phytate phosphorus by chicks. J. Nutr. 101:1289–1294. 10. Ravindran, V., E. T. Kornegay, Z. Yi, D. M. Denbow, and R. M. Hulet. 1995. Response of turkey poults to tiered levels of Natuphos phytase added to soybean meal-based semi-purified diets containing three levels of nonphytate phosphorus. Poult. Sci. 74:1843–1854. 11. Qian, H., E. T. Kornegay, and D. M. Denbow. 1996. Phosphorus equivalence of microbial phytase in turkey diets as influenced by calcium and phosphorus ratios and phosphorus levels. Poult. Sci. 75:69–81. 12. Kies, A. K., K. H. F. Van Hermert, and W. C. Sauer. 2001. Effects of phytase on protein and amino acid digestibility and energy utilization. World’s Poult. Sci. J. 57:109–126. 13. Greiner, R., U. Konietzny, and K. D. Jany. 1993. Purification and characterization of two phytases from Escherichia coli. Arch. Biochem. Biophys. 303:107–113. 14. Wodzinski, R. J., and A. H. Ullah. 1996. Phytase. Adv. Appl. Microbiol. 42:263–302. 15. Igbasan, F. A., K. Manner, G. Miksch, R. Borriss, A. Farouk, and O. Simon. 2000. Comparative studies on the in vitro properties

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

Ca and percentage of total P retention (r = 0.372, P = 0.0152), and between total P and percentage of phytate P disappearance (r = 0.464, P = 0.002). The significant correlations in chicks fed no enzyme were found between total P and percentage of total P retention (r = 0.355, P = 0.0287) and between protein and percentage of total P retention (r = 0.402, P = 0.0124). In Experiment 2, a significant correlation for chicks fed phytase was found only between total P and percentage of total P retention (r = 0.319, P = 0.0166). A significant correlation in Experiment 2 for chick groups fed soybean diets without phytase was found only

305

JAPR: Research Report

306 of phytases from various microbial origins. Arch. Tierernahr. 53:353–373. 16. Igbasan, F. A., O. Simon, G. Miksch, and K. Manner. 2001. The effectiveness of an Escherichia coli phytase in improving phosphorus and calcium bioavailability in poultry and young pigs. Arch. Tierernahr. 54:117–126. 17. Nelson, T. S., L. W. Ferrara, and N. L. Storer. 1968. Phytate phosphorus content of feed ingredients derived from plants. Poult. Sci. 47:1372–1374. 18. Denbow, D. M., V. Ravindran, E. T. Kornegay, Z. Yi, and R. M. Hulet. 1995. Improving phosphorus availability in soybean meal for broilers by supplemental phytase. Poult. Sci. 74:1831–1842.

20. Mitchell, R. D., and H. M. Edwards, Jr. 1996. Additive effects of 1,25-dihydroxycholicalciferol and phytase on phytate phosphorus utilization and related parameters in broiler chicks. Poult. Sci. 75:111–119. 21. Kornegay, E. T., D. M. Denbow, Z. Yi, and V. Ravindran. 1996. Response of broilers to graded levels of microbial phytase added to maize-soyabean meal based diets containing three levels of nonphytate phosphorus. Br. J. Nutr. 75:839–852. 22. Simons, P. C. M., H. A. J. Versteegh, A. W. Jongbloed, P. A. Kemme, P. Slump, K. D. Bos, M. G. E. Wolters, R. F. Beudeker, and G. J. Verschoor. 1990. Improvement of phosphorus availability by microbial phytase in broilers and pigs. Br. J. Nutr. 64:525–540. 23. Keshavarz, K. 2000. Reevaluation of nonphytate phosphorus requirement of growing pullets with and without phytase. Poult. Sci. 79:1143–1153. 24. Shirley, R. B., and H. M. Edwards, Jr. 2003. Graded levels of phytase past industry standards improves broiler performance. Poult. Sci. 82:671–680. 25. Leske, K. L., and C. N. Coon. 2002. The development of feedstuff retainable phosphorus values for broilers. Poult. Sci. 81:1681–1693. 26. Cobb 500, Cobb-Vantress Inc., Siloam Springs, AR. 27. Harlan Teklad, Madison, WI. 28. Araba, M., and N. M. Dale. 1990. Evaluation of protein solubility as an indicator of over processing soybean meal. Poult. Sci. 69:76–83. 29. American Association of Cereal Chemists. 2000. Approved Methods of the American Association of Cereal Chemists. 10th ed. Am. Assoc. Cereal Chem., St. Paul, MN. 30. National Research Council. 1994. Nutrient Requirements of Poultry. 9th rev. ed. Natl. Acad. Press, Washington, DC. 31. Celite Corp., Lompoc, CA. 32. Phyzyme XP 5000G is a microbial phytase enzyme preparation from Danisco Animal Nutrition (Marlborough, UK). Phyzyme XP 5000G phytase from E. coli expressed in yeast is a 6-phytase with acid phosphatase activity (with pH optimum of 2 to 5.5) that initiates hydrolysis at position 6 of the inositol-6-phosphate molecule when the pH is at the upper acidic range and the same phytase molecule has acid phosphatase activity when the pH is more acidic. 5000 refers to the guaranteed minimum enzyme activity in phytase units (FTU) per gram and G refers to the product form (i.e., dry and 1 FTU is the amount of enzyme that liberates 1 ␮mol of inorganic phosphate per minute from sodium phytate at pH 5.5 and 37°C. 33. Scott, T. A., and D. Balnave. 1991. Influence of temperature, dietary energy, nutrient concentration and self-selection feeding on the retention of dietary energy, protein and calcium by sexually maturing egg-laying pullets. Br. Poult. Sci. 32:1005–1016. 34. Bos, K. D., C. Verbeek, C. H. P. van Eeden, P. Slump, and M. G. E. Wolters. 1991. Improved determination of phytate by ion exchange chromatography. J. Agric. Food Chem. 39:1770–1772.

36. ANKOM Technol. Corp., Fairport, NY. 37. SAS Institute. 1999. SAS/STAT User’s Guide. Version 8. Vol. 1 and 2. SAS Institute Inc., Cary, NC. 38. Perney, K. M., A. H. Cantor, M. L. Straw, and K. L. Herkelman. 1993. The effect of dietary phytase on growth performance and phosphorus utilization of broiler chickens. Poult. Sci. 72:2106–2114. 39. Sebastian, S., S. P. Touchburn, E. R. Chavez, and P. C. Lague. 1996. Efficacy of supplemental phytase at different dietary calcium levels on growth performance and mineral utilization of broiler chickens. Poult. Sci. 75:1516–1523. 40. Qian, H., E. T. Kornegay, and H. P. Veit. 1996. Effects of supplemental phytase and phosphorus on histological, mechanical and chemical traits of tibia and performance of turkeys fed on a soybeanmeal-based semi-purified diets high in phytate phosphorus. Br. J. Nutr. 76:263–272. 41. Rama Rao, S. V., V. Ravindra Reddy, and V. Ramasubba Reddy. 1999. Enhancement of phytate phosphorus availability in the diets of commercial broilers and layers. Anim. Feed Sci. Technol. 79:211–222. 42. Ahmad, T., S. Rasool, M. Sarwar, A. Haq, and Z. Hasan. 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. 43. Viveros, A., A. Brenes, I. Arija, and C. Centeno. 2002. Effects of microbial phytase supplementation on mineral utilization and serum enzyme activities in broiler chicks fed different levels of phosphorus. Poult. Sci. 81:1172–1183. 44. Knuckles, B. E., and A. A. Betschart. 1987. Effect of phytate and other myo-inositol phosphate esters on alpha-amylase digestion of starch. J. Food Sci. 52:719–721. 45. Selle, P. H., V. Ravindran, R. A. Caldwell, and W. L. Bryden. 2000. Phytate and phytase: Consequences from protein utilization. Nutr. Res. Rev. 13:255–278. 46. Qian, H., E. T. Kornegay, and D. M. Denbow. 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. 47. Broz, J., P. Oldale, A. H. Perrin-Voltz, G. Rychen, J. Schulze, and C. S. Nunes. 1994. Effects of supplemental phytase on performance and phosphorus utilization in broiler chickens fed a low phosphorus diet without addition of inorganic phosphates. Br. Poult. Sci. 35:273–280. 48. Ravindran, V., S. Cabahug, G. Ravindran, P. H. Selle, and W. L. Bryden. 2000. Response of broiler chickens to microbial phytase supplementation as influenced by dietary phytic acid and non-phytate phosphorus levels. II. Effects on apparent metabolisable energy, nutrient digestibility and nutrient retention. Br. Poult. Sci. 41:193–200. 49. Edwards, H. M., Jr. 1993. Dietary 1,25-dihydroxycholecalciferol supplementation increases natural phytate phosphorus utilization in chickens. J. Nutr. 123:567–577. 50. Biehl, R. R., and D. H. Baker. 1997. Microbial phytase improves amino acid utilization in young chicks fed diets based on soybean meal but not diets based on peanut meal. Poult. Sci. 76:355–360. 51. Coon, C. N., and M. K. Manangi. 2004. The effect of a novel phytase on retainable phosphorus in broilers. Pages 56–68 in Proceedings of the 2nd Mid-Atlantic Nutr. Conf. N. G. Zimmermann, ed., Univ. Maryland, College Park.

Acknowledgments The authors are indebted to Danisco Animal Nutrition (Marlborough, UK) for financially supporting this research related to the phytase effect on total P retention and phytate P disappearance.

Downloaded from http://japr.oxfordjournals.org/ at East Tennessee State University on June 6, 2015

19. Scho¨ner, F. J., P. P. Hoppe, and G. Schwarz. 1991. Comparative effects of microbial phytase and inorganic phosphorus on performance and on retention of phosphorus, calcium and crude ash in broilers. J. Anim. Physiol. Anim. Nutr. 66:248–255.

35. Association of Official Analytical Chemists. 1990. Official Methods of Analysis. 15th ed. Assoc. Off. Anal. Chem., Washington, DC.