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Standardized Ileal Amino Acid Digestibility of Meat and Bone Meal from Different Sources in Broiler Chicks and Turkey Poults with a Nitrogen-Free or Casein Diet¹

S. A. Adedokun^*, C. M. Parsons, M. S. Lilburn, O. Adeola^* and T. J. Applegate^*,2

^* Department of Animal Sciences, Purdue University, 915 W. State Street, West Lafayette, IN 47907-2054; Department of Animal Sciences, University of Illinois, Urbana-Champaign 61801; and Department of Animal Sciences, The Ohio State University, Ohio Agricultural Research and Development Center, Wooster 44691

² Corresponding author: applegt{at}purdue.edu

	ABSTRACT

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The aim of this study was to determine the effect of the standardization method and age on standardized ileal amino acid digestibility (SIAAD) of 4 meat and bone meal (MBM) samples in broiler chicks and turkey poults. The SIAAD were obtained by correcting apparent ileal digestibility (AID) values for basal ileal endogenous amino acid flow obtained from chicks and poults fed a N-free diet or a highly digestible protein diet (HDP; 100 g of casein/kg of diet). Four diets were formulated to contain 20% CP by using MBM as the only source of CP. Each diet was fed to 6 replicate cages of 30 or 8 birds/cage at 5 and 21 d, respectively, for 5 d. With the exception of all-beef MBM, chick AID values on d 21 were higher (P < 0.05) than on d 5. In poults, however, AID was not different between the 2 ages, with the exception of His, which was higher (P < 0.05) at d 21. There was no difference between methods on SIAAD on d 5 or 21 for all 4 MBM samples in either species. There was no effect of the method of standardization on SIAAD at either age for most amino acids. For chicks, however, standardization of the all-beef MBM sample with HDP resulted in higher (P < 0.05) SIAAD values for most of the amino acids (d 5). In poults at d 5, however, the SIAAD for most of the amino acids in the blended MBM were higher (P < 0.05) with HDP correction. The CV in SIAAD in chicks between the 2 methods of standardization was higher on d 5 (8.1%) relative to d 21 (5.0%) for all treatments. In poults, greater variability was observed on d 21 (8.2%) relative to d 5 (6.7%). Results from this study showed that at d 21, SI-AAD with either the N-free or the HDP diet resulted in similar digestibility values.

Key Words: chick • ileal endogenous amino acid • meat and bone meal • poult • standardized ileal digestibility

	INTRODUCTION

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For optimal poultry production, it is important to formulate diets that meet the nutrient requirements for various classes of poultry while minimizing the excretion of excess nutrients into the environment. To achieve this goal, with respect to dietary protein and amino acids, accurate information on amino acid digestibility in common feed ingredients is required. Considerable data can be found in the literature on amino acid digestibility in poultry, but most of the published literature does not take into consideration the endogenous amino acid (EAA) contributions to ileal digesta or excreta. If diet formulations are based on apparent ileal digestibility (AID) values, the digestibility of amino acids in feedstuffs will be underestimated and may result in an increased level of nutrient excretion because of overformulation.

The contribution of ileal EAA (IEAA) to the digesta originates from various digestive secretions, including salivary, bile, pancreatic, gastric, and intestinal secretions. Other sources of EAA include mucoproteins and desquamated epithelial cells from the intestinal lining, as well as nondietary N originating from exocrine pancreatic secretions (Greene, et al., 1963; Nyachoti et al., 1997; Ravindran and Bryden, 1999). In correcting for basal IEAA secretions by using either the N-free diet (NFD) or highly digestible protein diet (HDP) method, the standardized ileal amino acid digestibility (SIAAD) values will be closer to the true digestibility value compared with the apparent digestibility value because of the presence of diet-specific EAA and the undigested portion of the diet in the digesta that is uncorrected for. A number of techniques have been used to estimate IEAA flow. Each of these techniques has strengths and weaknesses. According to Huang et al. (2005), the AID of CP and amino acids is affected by the age of the bird, but these effects varied among amino acids and ingredients.

A number of factors have been reported to contribute to the variability in apparent ileal amino acid digestibility values. These include the level of inclusion of the raw materials in the diet, the age of the birds (Ravindran and Hendriks, 2004), and the digestibility of the amino acids in the raw material itself (Lemme et al., 2004). Tarvid (1995) reported that digestion and absorption of dietary protein is influenced by age; however, there are contradictory reports on the effects of age on protein and amino acid digestibility in broiler chickens. Zuprizal and Chagneau (1992) reported a decrease in CP and amino acid digestibility of rapeseed meal with age (3 to 6 wk). Haakansson and Eriksson (1974) and Fonolla et al. (1981) also reported a decline in CP and amino acid digestibility of feed mixtures with age. These age effects may be a result of the contribution of amino acids of endogenous origin to the digesta. To take into account some of these factors that influence digestibility, diets are often formulated with a safety margin. This may contribute to increased N and amino acid excretion, which can increase environmental pollution as well as increase the cost of production.

Establishing SIAAD values at different ages for a variety of ingredients used in poultry diets will enable nutritionists to formulate diets based on the digestible amino acid content of the ingredient rather than on its total amino acid (TAA) concentration. This can potentially lead to a reduction in safety margins and a parallel reduction in N and amino acid excretion. The rationale for formulating diets based on digestible amino acids has been reviewed by Lemme et al. (2004).

The objective of this study was to determine the AID and SIAAD in broiler chicks and turkey poults at 2 ages (d 5 and 21) by using the NFD and HDP methods to correct for basal IEAA flow. In addition, the effects of age on SIAAD in broiler chicks and turkey poults were determined. Complete digestion and absorption of amino acids in casein were assumed in this study.

	MATERIALS AND METHODS

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Meat and Bone Meal
Four meat and bone meal (MBM) samples from different sources were used in this study. The MBM samples were from all beef (MBM1), all pork (MBM2), blended (MBM3), and blended plus grocery and packer trimmings (MBM4).

Diet Formulation
Two semipurified control diets were used for the determination of basal endogenous amino acid flow. These values were used to standardize AID to obtain SIAAD. The control diets were a NFD and a diet containing 100 g of casein/kg of diet (HDP). The remaining 4 diets were formulated to contain approximately 20% CP, with each of the 4 MBM samples supplying all the CP content in the diets. The composition and the analyzed amino acid composition of the experimental diets are reported in Tables 1 and 2, respectively. Two basal diets (a broiler starter and a turkey starter) were also made. Chromic oxide was added as the indigestible marker at a level of 3 g/kg of diet. Broiler and turkey poults were fed the same experimental diets, but the studies were conducted at 2 different locations.

Diets used for estimating IEAA and TAA flow were semipurified and contained 0 or 100 g of casein/kg of diet. Casein supplied all the amino acids in the latter diet. The control diets and the diets containing MBM were fed for 5 consecutive days before ileal contents were collected. On d 5, the birds in 6 replicate battery cages (n = 30 birds per cage) were euthanized and the ileal contents were removed by flushing with distilled water (chicks) or by gently squeezing the digesta (poults).

All remaining birds were fed a conventional corn- and soybean meal-based starter diet appropriate for the respective species until d 16. At this age, 288 birds from each species were randomly allocated to cages (n = 8 birds per cage). On d 21, all birds were euthanized and the ileal contents were collected as described for d 5. All birds were euthanized by CO₂ asphyxiation.

Birds were raised in battery cages (Alternative Design Manufacturing and Supply Inc., Siloam Springs, AR) until the end of the study in an environmentally controlled room with 24 h of light. Room temperature was 35°C during the first week and was dropped by 5°C during the subsequent weeks. Birds had free access to feed and water. All animal care procedures were approved by the Purdue University and the Ohio State University Animal Care and Use Committee.

Sampling and Ileal Digesta Processing
The ileum was considered to be that portion of the small intestine from Meckel’s diverticulum to approximately 5 mm proximal to the ileocecal junction. Ileal contents were flushed with distilled water by using 50-mL syringes (d 5) and wash bottles (d 21). The ileal digesta from all birds within a cage were pooled and stored in a freezer (–20°C) until they were processed. Samples were freeze-dried, ground with a mortar and pestle, and sent to the University of Missouri Experimental Station and Chemical Laboratories for complete amino acid and chromium analysis.

Chemical Analysis
Prior to diet formulation, the CP, N x 6.25 (%), ash (%), and peroxide values (mEq/kg; AOAC, 2000; method 957.08) of the MBM samples were determined on an as-is basis. The CP, ash, and peroxide contents of the MBM samples (air-dried basis) were 45.5, 33.5%, and 4.3 mEq/kg (MBM1); 55.5, 23.7%, and 109.3 mEq/kg (MBM2); 54.7, 22.1%, and 20.3 mEq/kg (MBM3); and 53.3, 24.2%, and 16.4 mEq/kg (MBM4).

Dry matter content was determined on ground diets, and ileal digesta was determined by drying the samples at 100°C for 24 h. Samples for amino acid analyses were hydrolyzed in 6 N HCl for 24 h at 110°C under a N atmosphere. For the sulfur-containing amino acids Met and Cys, performic acid oxidation was carried out before acid hydrolysis. Samples for Trp analysis were hydrolyzed by using barium hydroxide (AOAC, 2000; method 982.30 E [a, b, c]. The amino acids in the hydrolysate were subsequently determined by HPLC after post-column derivatization. Amino acid concentrations were not corrected for incomplete recovery resulting from hydrolysis. Chromium was determined by the inductively coupled plasma atomic emission spectroscopy method (AOAC, 2000; method 990.08) following nitric-perchloric acid wet ash digestion. Nitrogen in the MBM samples was determined by the combustion method (AOAC, 2000; method 990.03; model FP2000, Leco Corp., St. Joseph, MI) by using EDTA as a standard. Ash content was determined by drying the sample overnight at 100°C, followed by ashing in a muffle furnace for 18 h at 600°C.

Calculations
The ileal EAA and TAA flow from both species was calculated as milligrams of amino acid and TAA flow per kilogram of feed on a DM intake (DMI) basis by using the following formula (Moughan et al., 1992):

Standardized ileal amino acid digestibility (%) was calculated by using the formula given by Lemme et al. (2004):

Statistical Analysis
Data were analyzed by using the GLM procedure of SAS (SAS Institute, 2000) as a completely randomized design. The level of significance was set at P < 0.05.

	RESULTS

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For MBM2 and MBM3, the AID for DM and all the amino acids except for Trp and hydroxylysine were higher (P < 0.05) on d 21 compared with AID on d 5 (Table 3). For MBM4, AID for many of the amino acids was significantly higher at d 21 than at d 5. Apparent ileal digestibility values for TAA increased from d 5 to 21 by 17.5 (MBM2) and 17.7 (MBM3) percentage points and were not different for MBM4. However, AID for MBM1 decreased by 11 percentage points from d 5 to 21.

Of all the MBM samples used in this study, only His showed a significant improvement in digestibility with age in turkey poults (Table 6). The difference in digestibility between d 5 and 21 for His for MBM1, MBM2, MBM3, and MBM4 were 31.1, 42.3, 30.7, and 35.8 percentage points, respectively. The differences in TAA digestibility between d 5 and 21 were, respectively, 0.7, 4.0, 1.2 (a decrease), and 5.4 percentage points, and were not significant.

	DISCUSSION

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The objective of this study was to determine and compare the effects of standardization methods (NFD vs. HDP) and of age (d 5 vs. d 21) on AID and SIAAD values of 4 different MBM sources fed to broiler chicks and turkey poults. The experimental conditions and diets were similar and the health of the birds was excellent (total mortality was less than 2.5% for each of the treatments).

For the most accurate determination of amino acid digestibility in feed ingredients, it is essential to account for amino acids of endogenous origin in the digesta. Basal endogenous amino acids have been estimated in chickens by using NFD, HDP, regression methods, and fasted cockerels (Sibbald, 1979; Okumura et al., 1981; Ravindran and Bryden, 1999; Adedokun et al., 2007). However, dietary protein level affects EAA secretions, and ultimately the AID and SIAAD values. Correcting the AID of MBM for IEAA flow in 5-wk-old broilers has been reported to increase Lys (10.6%) and Met (7.1%) digestibility values (Angkanaporn et al., 1996).

The apparent ileal amino acid digestibility as determined in chicks increased in 2 (MBM2 and MBM3) of the 4 MBM samples for most amino acids, with the exception of Trp and hydroxylysine. With respect to MBM4, hydroxylysine and hydroxyproline did not show any improvement with age. In addition, only Ile, Leu, Met, Phe, Thr, Val, Ala, Glu, and Tyr showed improvement in digestibility with age. The greatest improvements (percentage points) between 5 and 21 d for Lys and Met, respectively, were seen in MBM2 (17 and 25) and MBM3 (16 and 23).The AID decreased with age for MBM1 in chicks. This observation is difficult to explain. It is interesting to note that the high level of peroxide in MBM2 appears not to have affected the AID. Overall, the AID for amino acids from all the MBM samples on d 21 were similar to the mean AID values for 19 MBM samples reported by Ravindran et al. (2002).

The AID values on d 21 were lower that those reported by Huang et al. (2005) in 14- and 21-d-old broilers chicks fed a diet containing 20% of CP from MBM, but the AID values for Lys, Met, Thr, and other amino acids in the current study were higher than those reported by Angkanaporn et al. (1996) for 5-wk-old broilers. The discrepancies in AID between published reports are most likely due to processing methods and the sources of raw material (Shirley and Parsons, 2000; Karakas et al., 2001).

Apparent ileal amino acid digestibility in poults, with the exception of His, was not influenced by age. With the exception of the MBM1 on d 5, AID in poults was higher than that of the chicks at both ages. The ileal digesta samples in poults were collected by gently squeezing the digesta from the ileum as compared with flushing with distilled water, as was the case for chicks. The difference in collection method may have had some effects on AID values. To avoid confounding effects of sloughed epithelial cells in digesta when digesta are squeezed from the ileum, we suggest that flushing with distilled water will greatly reduce this effect.

The overall amino acid AID was higher in poults than chicks for all the MBM except for the MBM1 sample, in which digestibility at d 5 for chicks was higher than for poults. This suggests that turkey poults may be better able to digest amino acids in MBM at both ages (d 5 and 21) than chicks. Histidine stood out from the remaining amino acids on d 5 in poults because digestibility was significantly lower (41%, mean of the 4 MBM samples) than on d 21. This digestibility of hydroxyproline and hydroxylysine was relatively higher in poults than in chicks, but unlike what we observed in poults, the digestibility for these 2 amino acids had high SD in chicks, which is indicative of the degree of variation in digestibility between species or across experimental locations. This shows that, in most cases, age and the level of hydroxyproline and hydroxylysine did not have a significant impact on amino acid digestibility in the MBM samples studied. Hydroxyproline from porcine MBM seems to have been better digested by poults than other sources of MBM used in this study.

Standardization of the AID values resulted in SIAAD values that were different for each amino acid in the MBM sources evaluated. However, unlike the results obtained by Angkanaporn et al. (1996) in their study, in which the standardized values for all the amino acids were similar (about 73%), the SIAAD values obtained in the current study were different for each of the amino acids. The difference between the 2 studies may be the result of the standardization method. Angkanaporn et al. (1996) used the homoarginine method, in which homoarginine digestibility is considered as being representative of the standardized values for all amino acids within an ingredient. In addition, the effect of age (5- and 21-d-old vs. 35-d-old broiler chicks) on ileal digestibility may have influenced the final result. For the broiler chicks, the HDP method resulted in significantly higher SIAAD values for Ile, Glu, and Ser for all the MBM samples in chicks and higher Ile and Glu for MBM2 in poults. This shows that SIAAD for these amino acids in MBM will be method sensitive.

For most of the amino acids, this study shows that within a given age, the NFD and HDP methods could be used for standardization. Between ages (d 5 vs. d 21), however, there was a decrease in the difference in SIAAD between the 2 methods, from 4.3 (d 5, NFD) to 2.6 (d 21, HDP) percentage units for chicks and from 4.2 (d 5, NFD) to 5.7 (d 21, HDP) percentage units for poults. The higher values from the HDP method (poults) could be a result of increased secretion from the gastrointestinal tract in response to the dietary protein. This study shows that either of the 2 methods of standardization can be used for both chicks and poults within the first 21 d.

	ACKNOWLEDGMENTS

 
Partial funding for this project was supplied by the U.S. Poultry and Egg Association (Tucker, GA), Fats and Proteins Research Foundation (Alexandria, VA), Degussa Corporation (Kennesaw, GA), ADM Inc. (Decatur, IL), Novus International Inc. (St. Louis, MO), and Ajinomoto Heartland LLC (Chicago, IL).

	FOOTNOTES

 
¹ Journal Paper No. 2007-18119 of the Purdue University Agricultural Research Programs.

Received for publication April 20, 2007. Accepted for publication July 31, 2007.

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