Jennifer E. Wubben 1, David M. Albin 1, Michelle R. Smiricky 2 and Vince M. Gabert 3. Illinois at Urbana-Champaign. Introduction

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1 Ileal Digestibilities of Amino Acids in Growing Pigs fed Soybean Meal and Corn Starch-Based Diets Supplemented with Fiber-Degrading Enzymes and Steeping Jennifer E. Wubben 1, David M. Albin 1, Michelle R. Smiricky 2 and Vince M. Gabert 3 1 M. S. student; 2 Ph. D. student; 3 Assistant Professor; Department of Animal Sciences, University of Illinois at Urbana-Champaign Introduction The digestibility of amino acids and other nutrients by the pig is an important measure of efficiency when evaluating swine diets or ingredients used in swine diets. Any method of feeding or use of a feed additive that significantly increases the ileal digestibility of amino acids could be used to increase the efficiency of amino acid use by the pig, and decrease waste and nutrient losses from swine production. Feed enzymes have been used in an attempt to increase nutrient digestibility in swine (Bedford and Schulze, 1998). Much of the research with fiber-degrading enzymes has been conducted with betaglucanase (Baas and Thacker, 1996; Li et al., 1996; Baidoo et al., 1998), which is commonly used in diets that mainly consist of barley or wheat. Research conducted with cellulase and hemicellulase is less common, and the influence of these two enzymes used in combination has not been evaluated. Soybean meal contains 3.39% cellulose and 3.48% hemicellulose on a dry matter basis (Gabert, 1994). Fibrous components of the diet, especially cellulose, have been shown to decrease amino acid digestibility in the pig (Li et al., 1994; Lien, 1995). Thus, cellulase and hemicellulase may improve the ileal digestibility of amino acids in soybean meal. Feed steeping (soaking meals in water prior to feeding) has been evaluated, and has been shown to increase weight gain and feed efficiency, as well as the efficacy of supplemental phytase and its effects on phosphorus retention, in a corn-soybean meal diet for growing pigs (Liu et al., 1997). Skoglund et al. (1997) also reported improvements in phosphorus utilization when a barleyrapeseed cake-pea diet was steeped. However, Nasi and Helander (1994), while conducting research with phytase, reported that soaking did not affect the digestibility or retention of any nutrient in a barley-soybean meal diet. The purpose of the present study was to evaluate the effect of cellulase and hemicellulase, feed steeping for 12 h prior to feeding, and the combination of both of these treatments, on ileal amino acid digestibilities in growing pigs fed semipurified diets with soybean meal as the sole protein source. Materials and Methods

2 Animals and treatments. Eight barrows (Line 326 sire x C22 dams; PIC, Franklin, KY) with an average initial body weight of 69.9 kg were surgically fitted with a simple-t cannula at the distal ileum using procedures adapted from Sauer et al. (1983). The barrows were housed in individual metabolism crates in a temperature-controlled barn, and were used for two experimental periods. Two experimental diets with (Diet B) and without (Diet A) fiber-degrading enzymes, formulated to contain 17% crude protein and.5% chromic oxide, were used (Table 1). The enzyme diet (B) contained 30,080 units of cellulase/kg diet and 531 units of hemicellulase/kg diet. The barrows were fed one of four experimental treatments control diet (A); control diet plus steeping for 12 h prior to feeding; enzyme diet (B); and the enzyme diet plus steeping for 12 h prior to feeding. Water was added to the steeped meals on a 1:1 weight basis, and was available to all pigs from a low-pressure drinking nipple ad libitum. Each period consisted of a 5 d adaptation phase and 2 d collection phase. The pigs were fed 1.76 kg/d in the first period, and 1.84 kg/d in the second period. Feeding occurred at 0800 and 2000 h. Ileal digesta were collected for 12 h intervals, from 0800 to 2000 h, over the two days. Digesta was frozen at 10 o C after collection, and was pooled within barrow during the first period. During the second period, digesta was pooled within barrow and day of collection to determine the effect of day on digestibility calculations. Chemical and statistical analysis. Samples of diets, freeze-dried digesta and soybean meal were ground with liquid nitrogen in a coffee mill. For dry matter determinations, empty 50 ml glass beakers that had been in a 105 o C oven overnight were weighed immediately after removal from the oven. Samples of diets and digesta were then weighed into the beakers, and the beakers plus sample were placed into the oven overnight. The next day each beaker and sample were weighed, and dry matter was calculated. The same samples were then ashed in a muffle furnace for 16 h at 450 o C. Organic matter was determined by the difference between ash percentage and 100. Then, the samples were used for chromic oxide determination according to Fenton and Fenton (1979). Samples of diets and digesta were analyzed for crude protein according to the AOAC (1995). Amino acid levels were determined in the diets, digesta, soybean meal, cellulase and hemicellulase using pre-column derivatization with phenyl isothiocyanate. Samples were weighed into screwcapped test tubes, and 6 M HCl was added to the tubes. The tubes were purged with nitrogen and were hydrolyzed in an oven for 24 h at 110 o C. Amino acids were determined in our laboratory with procedures adapted from Bidlingmeyer et al. (1984). Digesta, cellulase and hemicellulase were analyzed in duplicate, soybean meal in triplicate, and five samples of each diet were analyzed. Correction factors determined in our laboratory were used to more accurately reflect amino acid concentrations in both diets and digesta. In the second period, day was found to have no influence (P >.05) on apparent ileal digestibility. Therefore, digestibility coefficients for day 1 and day 2 were not pooled, and thus, six replicates per treatment were obtained. Also, the use of correction factors did not affect (P >.05) digestibility determinations when compared to those determined without correction factors. The apparent ileal digestibility values of the experiment treatments were subjected to statistical analysis as a twoperiod crossover design with six replicates per treatment using PROC GLM of SAS (1990). Means of dietary treatments and experimental periods were compared using Fisher s protected least significant difference procedure (pdiff procedure, SAS; Milliken and Johnson, 1984). Results and Discussion The pigs remained healthy and consumed their daily meals throughout the experiment.

3 The apparent ileal digestibilities of dry matter, crude protein, organic matter and amino acids are presented in Table 2. Digestibility was not affected (P >.05) by any of the treatments. There were small numerical increases in digestibility with the use of enzymes, as well as with the use of steeping. The use of fiber-degrading enzymes in conjunction with steeping numerically reduced the digestibility of some amino acids. To our knowledge, this is the first study to investigate the influence of a combination of fiber-degrading enzymes on the ileal digestibility of amino acids. Our findings with regard to steeping are in agreement with those of Nasi and Helander (1994). Implications Fiber-degrading enzymes and steeping did not affect ileal amino acid digestibility in growing pigs. Further research with pigs at different ages and physiological states, as well as different diets, should be conducted to further evaluate the efficacy of fiber-degrading enzymes and steeping. Literature Cited AOAC Official Methods of Analysis (16 th Ed.). Association of Official Analytical Chemists, Arlington, VA. Baas, T.C., and P.A. Thacker Impact of gastric ph on dietary enzyme activity and survivability in swine fed b-glucanase supplemented diets. Can. J. Anim. Sci. 76: Baidoo, S.K., Y.G Liu, and D. Yungblut Effect of microbial enzyme supplementation on energy, amino acid digestibility and performance of pigs fed hulless barley based diets. Can. J. Anim. Sci. 78: Bedford, M.R., and H. Schulze Exogenous enzymes for pigs and poultry. Nutr. Res. Rev. 11: Bidlingmeyer, B.A., S.A. Cohen, and T.L. Tarvin Rapid analysis of amino acids using pre-column derivatization. J. Chromatogr. 336: Fenton, T.W., and M. Fenton An improved procedure for the determination of chromic oxide in feed and feces. Can. J. Anim. Sci. 76: Gabert, V.M The effect of supplementation of organic acids, oligosaccharides and lactitol on the digestibility of amino acids and bacterial populations and metabolites in the digestive tract of early-weaned pigs. M.S. thesis. University of Alberta, Edmonton. Li, S., W.C. Sauer, and R.T. Hardin Effect of dietary fibre level on amino acid digestibility in young pigs. Can. J. Anim. Sci. 74: Li, S., W.C. Sauer, and S.X. Huang Effect of b-glucanase supplementation to hulless barley- or wheat-soybean meal diets on the digestibilities of energy, protein, b-glucans, and amino acids in young pigs. J. Anim. Sci. 74:

4 Lien, K.A Endogenous protein in ileal digesta: estimation of the mucin content and an evaluation of the 15 N-isotope dilution technique. Ph.D. thesis. University of Alberta, Edmonton. Liu, J., D.W. Bollinger, and D.R. Ledoux Soaking increases the efficacy of supplemental phytase in a low-phosphorus corn-soybean meal diet for growing pigs. J. Anim. Sci. 75: Milliken, G.A. and D.E. Johnson Analysis of Messy Data Volume 1: Designed Experiments. Van Nostrand Reinhold, NewYork. Nasi, M., and E. Helander Effects of microbial phytase supplementation and soaking of barley soybean meal on availability of plant phosphorus for growing pigs. Acta. Agric. Scand. Sect. A. 44: SAS SAS/STAT User s Guide (Release 6.08). SAS Inst. Inc., Cary, NC. Sauer, W.C., H. Jorgensen, and R. Berzins A modified nylon bag technique for determining apparent digestibilities of protein in feedstuffs for pigs. Can. J. Anim. Sci. 63: Skoglund, E., T. Larsen, and A.S. Sandberg Comparison between steeping and pelleting a mixed diet at different calcium levels on phytate degradation in pigs. Can. J. Anim. Sci. 77: Table 1. Formulation (%) of the experimental diets (as-fed basis) Diet Ingredient A B Corn starch a Soybean meal Sucrose b Corn oil Dicalcium phosphate Calcium carbonate Vitamin premix c Trace mineral premix d Cellulase e Hemicellulase f Chromic oxide a Clinton Brand, Archer Daniels Midland Company, Effingham, IL b Domino Sugar, Tate & Lyle North American Sugars, Inc., New York, NY c Provided the following (per kg of diet): choline, 121 mg; niacin, 16.5 mg; pantothenic acid, 12.1 mg; riboflavin, 4.40 mg; vitamin B 12,.020 mg; vitamin E, 44.1 IU; vitamin A, 3310 IU; vitamin D, 331 IU; vitamin K, 2.21 mg. Supplied by Archer Daniels Midland Company, Effingham, IL.

5 d Provided the following (per kg of diet): sodium, 2.80 g; chloride, 2.80 g; copper, 7.79 mg; iodine,.34 mg; iron, 87.4 mg; manganese, 19.4 mg; selenium, 0.29 mg; zinc, 97.2 mg. Supplied by Archer Daniels Midland Company, Effingham, IL. e Cellulase, Sigma Chemical Company, cat. # C-0901, provided 30,080 enzyme units per kg diet. kg diet. f Hemicellulase, Sigma Chemical Company, cat. # H-2125, provided 531 enzyme units per Table 2. Apparent ileal digestibilities (%) of dry matter, organic matter, crude protein and amino acids in the experimental treatments Treatment a Item A A plus steeping B B plus steeping SEM b Dry matter Organic matter Crude protein Amino acids c Indispensable Arginine Histidine Isoleucine Leucine Lysine Phenylalanine Threonine Valine Dispensable Alanine Aspartic acid Glutamic acid Glycine Serine Tyrosine Proline a Treatment did not affect (P >.05) apparent ileal digestibility. b Standard error of the mean (n=6). c Amino acid digestibility coefficients calculated using correction factors determined in our laboratory. See Materials and Methods.