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Chemical composition, protein quality, palatability, and digestibility of alternative protein sources for dogs J. M. Dust*, C. M. Grieshop*, C. M. Parsons*, L. K. Karr-Lilienthal*, C. S. Schasteen†, J. D. Quigley, III‡, N. R. Merchen*, and G. C. Fahey, Jr.*1 *University of Illinois at Urbana-Champaign, Urbana 61801; †Novus International, Inc., St. Charles, MO 63304; and ‡APC, Ankeny, IA 50021 ABSTRACT: The chemical composition and protein quality of 11 alternative protein sources (chicken products, blood products, enzyme-hydrolyzed fish protein concentrate, soybean meal, and spray-dried pork liver) were determined, and an experiment was conducted to determine palatability and digestibility of processed red blood cell-containing diets. Chicken protein sources differed in concentrations of CP, acid-hydrolyzed fat, and total AA (TAA) by 20, 31, and 24%, respectively, and GE by 1.7 kcal/g. Blood protein sources varied little in acid-hydrolyzed fat and GE concentrations, but concentrations of CP and TAA differed by 11 and 8%, respectively. Protein solubility of chicken and blood protein source categories averaged 57 and 69%, respectively. Protein solubility of enzyme-hydrolyzed fish protein concentrate, soybean meal, and spray-dried pork liver was 53, 67, and 26%, respectively. Based on calculations from immobilized digestive enzyme assay values, lysine digestibility averaged approximately 80.4 and 81.7% for blood and chicken protein sources, respectively. Lysine digestibility values for soybean meal and spray-dried pork liver were 89 and 77%, respectively. A chick protein efficiency ratio (PER) assay showed that chicken protein sources had high protein quality values, as the PER ranged from 2.7 to 5.3, whereas blood protein sources had poor protein quality (PER values less than 1.5). Enzyme-hydrolyzed fish protein concentrate, spray-dried pork liver, and soybean meal had high protein quality (PER values greater than 2.8). In the dog palatability and digestibility experiments, a corn and chicken-based diet supplemented with either 0 or 3% processed red blood cells was tested. The palatability test showed that dogs consumed more of the diet that contained 0% vs. 3% processed red blood cells. The intake ratio for the 3% processed red blood cells diet was 0.34. Nutrient digestibilities did not differ, except for CP, where the digestibility was greater (P = 0.01) for dogs consuming the 0% processed red blood cells diet. These data suggest that chemical composition and quality of alternative protein sources differ greatly among ingredients within the same category. Palatability data suggest that a processed red blood cells-containing diet is not highly palatable but, when this diet was offered as only one food, dogs demonstrated no aversion response but some decrease in protein digestion. Key Words: Digestibility, Dog, Palatability, Protein Efficiency Ratio, Protein Quality, Protein Sources 2005 American Society of Animal Science. All rights reserved. Introduction A key component of canine diets is the protein source used. Protein contains essential AA as well as nonessential AA that are used for energy, muscle deposition, and metabolic functions of the body. The quantity of protein required by a dog is age-dependent (AAFCO, 2003). A puppy has a greater demand for protein, as its body is 1 Correspondence: 166 Animal Sciences Laboratory, 1207 W. Gregory Dr. (phone: 217-333-2361; fax: 217-244-3169; e-mail: gcfahey@ express.cites.uiuc.edu). Received April 29, 2004. Accepted June 20, 2005. J. Anim. Sci. 2005. 83:2414–2422 growing rapidly and a large amount of muscle is being deposited. In contrast, an adult dog requires less protein to maintain muscle mass and body metabolism (NRC, 1985). Today’s pet food industry is growing rapidly, with pet owners demanding high-quality diets for their pets. This demand is creating a search for alternative protein sources that may be included in diets to meet the AA requirements of the animal, to serve as palatants, and/ or to enhance the immune status of the animal. The pet food industry traditionally has used a wide range of protein sources, including meat and bone meals, poultry meals, poultry by-product meals, and soybean meal (AAFCO, 2003). Alternative protein sources that have been studied in other animal species, but only to a 2414 Alternative protein sources for dogs limited extent in companion animals, include spraydried plasma (Quigley et al., 2004), wheat protein, fish protein, and processed red blood cells. By expanding the database on compositional analyses of alternative protein sources and by determining their quality, their ability to enhance palatability, and their immunomodulatory role, nutritionists will be able to use these alternative ingredients in dietary formulations for pets to optimal advantage. Therefore, the objective of this study was to determine the chemical composition and protein quality of several alternative protein sources. In addition, an experiment to determine the palatability and digestibility of one of these sources, processed red blood cells, as a component of dog diets was conducted. Materials and Methods Substrates Eleven spray-dried or conventional protein sources were tested. Chicken protein sources included spraydried cooked chicken from deboned USDA-inspected chicken parts used for pet food applications that is then spray dried to form a powder (American Dehydrated Foods, Inc., Springfield, MO); spray-dried cooked chicken liver produced from USDA-inspected facilities using chicken livers that are ground, cooked, and spraydried (American Dehydrated Foods, Inc.); spray-dried egg processed from pasteurized whole egg solids, and then spray dried to form a granulated powder (American Dehydrated Foods, Inc.); chicken-by-product meal (Tyson Foods, Inc., Springdale, AR) comprising ground, cleaned, rendered carcass parts of chicken with trace amounts of feathers and blood (AAFCO, 2003); and poultry by-product meal (Tyson Foods, Inc.) comprising ground, cleaned, rendered carcass parts of poultry including heads, feet, and viscera, to include trace amounts of feathers and blood (AAFCO, 2003). The blood protein sources included processed red blood cells (American Protein Corp., Inc., Ankeny, IA) obtained from blood collected from abattoirs and centrifuged to separate the red blood cells from the plasma. The red blood cells are then processed using a proprietary procedure to produce a cream colored powder. Spray-dried plasma (American Protein Corp., Inc.) is a product of blood collected from abattoirs and centrifuged to separate the plasma fraction from the red blood cells. The plasma is then spray dried, producing a powder. Spray-dried whole beef blood (California Spray Dry Co., Stockton, CA) is whole blood with approximately 20% solids that is spray dried to form a free-flowing powder. Other protein sources tested included enzyme-hydrolyzed fish protein concentrate (California Spray Dry Co.), which is fish remaining after filleting that are then subjected to controlled proteolytic enzyme digestion that hydrolyzes the CP into peptides and free AA. Soybean meal (SBM) is a common plant protein source 2415 produced by removing the oil from soybeans and grinding the residue into a meal form. Spray-dried pork liver (California Spray Dry Co.) is pork liver from USDAinspected facilities that undergoes a solubilization process at a regulated temperature, time, and pH, followed by enzymatic hydrolysis, pasteurization, and spray drying. The SBM was ground with dry ice to pass 2- and 0.5mm screens in a model 4 Wiley mill (Thomas-Wiley, Swedesboro, NJ) in preparation for chemical analyses. Other protein products already were finely ground and required no further processing. Chemical Analyses Each protein source was analyzed for DM, OM, ash (AOAC, 1985), CP (AOAC, 1995) using a Leco nitrogen/ protein determinator (model FP-2000; Leco Corp., St. Joseph, MI), acid-hydrolyzed fat (Budde, 1952; AACC, 1983), and GE (Parr Instrument Manuals, Parr Instrument Co., Moline, IL). The 11 protein sources were analyzed for AA concentrations using a Beckman 6300 AA analyzer (Beckman Coulter, Inc., Fullerton, CA, according to AOAC, 1995) at the University of Missouri Experiment Station Chemical Laboratories. The reported AA concentrations were not corrected for incomplete recovery resulting from hydrolysis. All ingredients were analyzed in duplicate to provide a snapshot of protein availability by in vitro methods. Analyses were repeated if deviation of greater than 5% occurred among duplicates for all components present at a concentration greater or equal to 10%. Analyses were repeated if a deviation of greater than 10% occurred among duplicates for all components present at a concentration of less than 10%. Protein Quality Assays A protein solubility in potassium hydroxide assay (Araba and Dale, 1990) and the immobilized digestive enzyme assay (IDEA; Schasteen et al., 2002) were conducted on each protein source. A protein efficiency ratio (PER) assay was conducted utilizing 220, 8-d-old female chicks (New Hampshire × Columbian cross). The average initial BW was 91.8 g/chick. Chicks were housed in groups of five in starter batteries with raised wire floors in an environmentally regulated room. Eleven diets were formulated to contain one of the 11 protein sources in an otherwise N-free diet to provide 10% CP (Johnston and Coon, 1979). Composition of the diets is presented in Table 1. The experiment was a completely randomized design including 11 treatments, with four replications and five chicks per replication. The chicks were allowed ad libitum access to food and water over the 9-d assay period. Initial and final BW were recorded. Food consumption was recorded for calculation of G:F and PER. Data were analyzed as a completely randomized design using ANOVA in SAS (Version 8, SAS Inst., Inc., Cary, NC). Treatment differences 2416 Dust et al. Table 1. Chick protein efficiency ratio (PER) assay diet used to evaluate alternative protein sources Ingredient Cornstarch:dextrose (2:1 ratio) Protein sourcesa Soybean oil Limestone Dicalcium phosphate Sodium chloride MgSO4ⴢ7H2O K2CO3 Purified vitamin mixbc Mineral mixd Purified choline chloridec Ethoxyquin DL-Tocopherol acetate %, as-is basis To 100 To provide 10% CP 5.0 1.22 2.45 0.50 0.35 0.90 0.20 0.15 0.20 0.0125 0.002 a Protein sources (% incorporated into diet): chicken by-product meal (15.92%), poultry by-product meal (15.60%), spray-dried cooked chicken (20.35%), spray-dried cooked chicken liver (14.49%), spraydried egg product (18.96%), processed red blood cells (10.50%), spraydried plasma (11.85%), spray-dried whole beef blood (10.47%), enzyme-hydrolyzed fish protein concentrate (16.06%), soybean meal (19.63%), and spray-dried pork liver (14.36%). b Provided per kilogram of diet: thiamin, 20 mg; niacin, 50 mg; riboflavin, 10 mg; Ca-pantothenate, 30 mg; vitamin B12, 0.04 mg; pyridoxine, 6 mg; biotin, 0.6 mg; folic acid, 4 mg; vitamin K, 2 mg; vitamin D, 15 g; vitamin A, 1,789 g; vitamin C, 250 mg. c Choline chloride and vitamin mix in a carrier with little to no nitrogen-containing compounds were incorporated to provide an otherwise nitrogen-free diet except for nitrogen from test ingredients. d Provided per kilogram of diet: Mn (as MnO), 75 mg; Fe (as FeSO4∋H2O), 75 mg; Zn (as ZnO), 75 mg; Cu (as CuSO4ⴢ5H2O), 8 mg; I (as CaI2), 0.75 mg; Se (as Na2SeO3), 0.3 mg. were determined using the LSD calculated from the pooled SEM from ANOVA. A probability of P < 0.05 was considered statistically significant. Palatability Experiment Twenty beagle dogs (seven males and 13 females) with BW ranging from 6.3 to 19.9 kg were used. Dogs were individually housed in indoor-outdoor pens measuring approximately 1.2 × 1.5 m indoors and 1.2 × 3.0 m outdoors at Kennelwood, Inc., Champaign, IL. Dogs had access to the outside area of the kennel once daily for an average of 2 to 4 h, depending on weather conditions. Two diets were formulated representing a cornand chicken-based canine diet supplemented with either 0 or 3% processed red blood cells. Composition of the diets is presented in Table 2. Diets were formulated to be isocaloric. Dietary ingredients were identical except for supplementation of processed red blood cells in place of corn gluten meal. Diets were an extruded kibble, with the processed red blood cells being incorporated into the mixture before extrusion. The experiment was designed as a two-bowl, free-choice test, the most common palatability test in the petfood industry (Griffen, 2003). This design results in the most reliable data (Hutton, 2002). Dogs were on test for 4 d. All dogs were allowed free access to water. Dogs were offered 500 g each of the 0% processed red blood cells diet and the 3% processed red blood cells diet in separate bowls Table 2. Ingredient composition of diets containing processed red blood cells (PRBC) for palatability and digestibility experiments with dogs (%, as-is basis) Dietary treatment Ingredient Corn, ground Poultry by-product meal Brewer’s rice Soybean meal, dehulled Corn gluten meal Processed red blood cells Beet pulp Poultry fat KCl NaCl Choline chloridea Vitamin premixb Mineral premixc Ethoxyquin 0% PRBC 3% PRBC 44.47 15.40 14.50 12.00 4.15 — 4.00 4.00 0.60 0.50 0.13 0.12 0.12 0.02 45.62 15.40 14.50 12.00 — 3.00 4.00 4.00 0.60 0.50 0.13 0.12 0.12 0.02 a Provided per kilogram of diet: choline, 780 mg. Provided per kilogram of diet: vitamin A, 14,970 IU; vitamin D3, 900 IU; vitamin E, 59.88 IU; vitamin K, 0.60 mg; thiamin, 11.98 mg; riboflavin, 9.58 mg; pantothenic acid, 17.96 mg; niacin, 44.91 mg; pyridoxine, 11.98 mg; biotin, 0.11 mg; folic acid, 0.72 mg; vitamin B12, 0.02 mg. c Provided per kilogram of diet: Mn (as MnSO4), 12 mg; Fe (as FeSO4), 90 mg; Cu (as CuSO4), 12 mg; Co (as CoSO4), 2.4 mg; Zn (as ZnSO4), 120 mg; I (as KI), 1.5 mg; Se (as Na2SeO3), 0.24 mg. b for 1 h daily. The placement of the bowls was alternated each day to eliminate any bowl placement bias by the dogs. First-choice and first-approach data were collected. At the end of the hour, any refused food was weighed to determine food intake of each diet. A sample was taken of each diet and frozen at −4°C for subsequent analyses. Diets were ground with dry ice to pass a 2mm screen in a model 4 Wiley mill (Thomas-Wiley) in preparation for chemical analyses. Samples of each diet and the processed red blood cells ingredient were taken and analyzed for DM, OM, ash, CP, acid-hydrolyzed fat, and GE as in chemical analysis section. The quantity of each diet consumed was calculated by subtracting food refusal from amount of food originally offered. Intake ratio (IR) was calculated by dividing the grams consumed of the 3% processed red blood cells diet by the total grams consumed of both diets. Corrected intake ratio was calculated as intake ratio minus 0.5, to indicate a diet preference. An intake ratio of 0.5 indicates no preference; therefore, the corrected intake ratio will detect whether there was a diet preference significantly different from zero. Data were analyzed using the Mixed procedure of SAS. The model contained the fixed effect of diet and the random effect of dog. A probability of P < 0.05 was considered statistically significant. Digestibility Experiment Fourteen beagle dogs that were 2 to 11 yr of age and ranged in BW from 6.8 to 15.5 kg were used. Dogs were individually housed in indoor-outdoor pens measuring Alternative protein sources for dogs approximately 1.2 × 1.5 m indoors and 1.2 × 3.0 m outdoors at Kennelwood, Inc. As in the palatablity experiment, dogs had access to the outside area of the kennel once daily for an average of 2 to 4 h, depending on weather conditions. Diets were the same as ones tested in the palatability experiment (Table 2). Dogs were fed 500 g of the appropriate diet once daily, and food intake was measured daily. Any food refusal during the digestibility trial was collected and weighed daily. Water was provided for ad libitum consumption throughout the study. Dogs participated in a 20-d (two 10-d periods) digestibility experiment, in which each dog received the control (Period 1, n = 6; Period 2, n = 8) and processed red blood cells (Period 1, n = 8; Period 2, n = 6)-containing diet in a crossover design. Within each period, dogs were allowed a 7-d diet adaptation phase, after which a 3-d total fecal collection was conducted for determination of nutrient digestibilities. A sample was taken of each diet and frozen at −4°C for subsequent analyses. Diets were ground with dry ice, to avoid loss of lipid, to pass a 2-mm screen in a model 4 Wiley mill (Thomas-Wiley) in preparation for chemical analyses. Samples of the processed red blood cells, food, and fecal samples were prepared and analyzed as in palatability trial. Feces were scored and weighed at the time of collection. Scoring was determined using the following system: 1 = hard, dry pellets – small, hard mass; 2 = hard, formed, dry stool – remains firm and soft; 3 = soft, formed moist – softer stool that retains shape; 4 = soft, unformed – stool assumes shape of container; and 5 = watery – liquid that can be poured. Feces were frozen at −4°C and composited by dog at the end of each collection. All fecal samples were dried at 57°C and ground to pass a 2-mm screen in a model 4 Wiley mill (Thomas-Wiley). Data were analyzed in a crossover design using the Mixed procedure of SAS. The model contained the fixed effects of diet and period and the random effect of dog. A probability level of P < 0.05 was considered statistically significant. Results and Discussion Dry matter, OM, CP, acid-hydrolyzed fat, GE, and AA concentrations of protein sources are presented in Tables 3, 4, and 5. Dry matter values for the chicken protein sources (Table 3) varied little. Organic matter concentrations ranged from 87 to 94%. Crude protein concentrations ranged from 49.2% (spray-dried cooked chicken) to 69.0% (spray-dried cooked chicken liver). Acid-hydrolyzed fat content varied from 18.3% (chicken-by product meal) to 49.5% (spray-dried cooked chicken). Gross energy content was lowest for chicken by-product meal and poultry by-product meal and highest for spray-dried cooked chicken. Chicken by-product meal, poultry by-product meal, and spray-dried egg were similar in total essential amino acid (TEAA) com- 2417 position. Spray-dried cooked chicken liver had the greatest TEAA and spray-dried cooked chicken the least. Spray-dried cooked chicken had lower arginine, leucine, and phenylalanine concentrations than the other chicken protein sources. Chicken by-product meal had the greatest total nonessential amino acid (TNEAA) concentration (39.2%) and spray-dried cooked chicken the least (22.0%). Chicken by-product meal and poultry by-product meal were highest in glycine, proline, and hydroxyproline content. This result is perhaps indicative of the connective tissue content of these by-products compared with the other chicken protein sources. Poultry by-product meal has been reported to have CP concentrations ranging from 61.9 to 74.5% (Johnson and Parsons, 1997; Murray et al., 1997; Clapper et al., 2001). The CP values for chicken-by product meal and poultry by-product meal used in the present study fall within this range, with values being closest to those reported by Johnson and Parsons (1997). Fat concentrations in the present study were greater than those reported by Murray et al. (1997) and Clapper et al. (2001). Batch variation is likely the reason for the variable fat concentrations. Even though CP values for ingredients used in the present study were lower than those reported by Murray et al. (1997) and Clapper et al. (2001), TEAA concentrations were similar. Overall, poultry byproduct meal and chicken-by product meal had higher CP and total amino acid (TAA) concentrations than spray-dried cooked chicken and spray-dried egg, which is likely due to the connective tissue content of the byproduct meals. Chicken by-product meal and poultry by-product meal were included in the present study as they are typically used in pet food manufacturing for their nutrient content and palatability (Heinicke, 2003). For the blood protein sources tested (Table 4), DM concentrations were greatest for processed red blood cells and spray-dried whole beef blood (average = 96.8%) and least for spray-dried plasma (93.2%). Organic matter, CP, and GE concentrations were least for spraydried plasma, whereas fat concentration was greatest. For processed red blood cells, the TAA concentration was much lower than the CP concentration, whereas for spray-dried plasma, the TAA value was nearly three percentage units higher. The variation in AA concentrations between spray-dried plasma and processed red blood cells compared with spray-dried whole beef blood could be a result of not correcting the values for incomplete recovery during hydrolyis. Spray-dried blood meals from bovine and porcine sources vary little in chemical composition (Kats et al., 1994; DeRouchey et al., 2002), with average DM and CP concentrations of approximately 92 and 89%, respectively. These values are lower than those noted in the present study for processed red blood cells and spraydried whole beef blood. The chemical composition of processed red blood cells in the present study and spraydried blood cells (DeRouchey et al., 2002) are similar. 2418 Dust et al. Table 3. Chemical composition of chicken protein sourcesa Item CBM PBM DM, % 94.9 97.8 SDCC SDCCL SDE 97.7 98.3 95.9 DM basis OM, % CP, % Acid-hydrolyzed fat, % GE, kcal/g 88.2 62.8 18.3 5.5 86.7 64.1 19.6 5.5 94.2 49.2 49.5 7.2 93.2 69.0 22.7 6.0 94.0 52.7 36.9 6.7 Amino acids, % Essential Arginine Histidine Isoleucine Leucine Lysine Methionine Phenylalanine Threonine Tryptophan Valine TEAAb TNEAAc TAAd 4.81 1.37 2.42 4.49 4.06 1.35 2.54 2.37 0.48 3.16 27.1 38.7 65.8 4.44 1.39 2.39 4.54 4.11 1.33 2.45 2.33 0.55 3.15 26.7 36.1 62.8 2.84 1.32 1.96 3.41 3.65 1.06 1.75 1.61 0.42 2.28 20.3 21.6 41.8 4.00 1.72 2.82 5.72 4.73 1.58 3.06 2.72 0.90 3.87 31.1 29.6 60.7 3.13 1.39 3.10 4.85 3.77 1.76 2.91 2.16 0.83 3.64 27.5 24.8 52.4 a CBM = chicken by-product meal; PBM = poultry by-product meal; SDCC = spray-dried cooked chicken; SDCCL = spray-dried cooked chicken liver; SDE = spray-dried egg product. b TEAA = total essential amino acids. c TNEAA = total nonessential amino acids. d TAA = total amino acids. Kerr et al. (1998) reported that spray-dried plasma had CP and TEAA concentrations of 82.9 and 39.5%, respectively, values that are similar to ours. A concern with including blood products is that few palatability studies using blood meal have been tested in canine diets; however, blood meals have been shown to be unpalatable Table 4. Chemical composition of blood protein sourcesa Table 5. Chemical composition of enzyme-hydrolyzed fish protein concentrate (EHFPC), soybean meal (SBM), and spray-dried pork liver (SDPL) Item PRBC DM, % 96.6 SDP SDWBB 93.2 96.9 DM basis OM, % CP, % Acid-hydrolyzed fat, % GE, kcal/g 92.8 95.3 1.6 5.4 90.9 84.4 2.9 5.1 94.8 95.5 2.1 5.5 Amino acids, % Essential Arginine Histidine Isoleucine Leucine Lysine Methionine Phenylalanine Threonine Tryptophan Valine TEAAb TNEAAc TAAd 3.60 5.90 0.30 13.47 6.25 0.62 6.42 2.63 0.39 8.87 48.5 39.0 87.5 4.90 2.83 3.01 8.39 7.52 0.71 4.83 4.53 1.40 5.64 43.8 43.4 87.2 3.62 5.92 0.39 13.32 9.23 1.19 6.95 4.20 1.45 8.44 54.8 40.0 94.8 a PRBC = processed red blood cells; SDP = spray-dried plasma; SDWBB = spray-dried whole beef blood. b TEAA = total essential amino acids. c TNEAA = total nonessential amino acids. d TAA = total amino acids. Item EHFPC SBM SDPL DM, % 94.7 94.4 96.1 OM, % CP, % Acid-hydrolyzed fat, % GE, kcal/g 90.4 62.3 27.3 6.1 92.6 51.0 4.5 4.7 89.9 69.7 14.5 5.5 Amino acids, % Essential Arginine Histidine Isoleucine Leucine Lysine Methionine Phenylalanine Threonine Tryptophan Valine TEAAa TNEAAb TAAc 2.92 1.23 2.46 3.97 3.99 1.26 2.10 2.08 0.49 3.18 23.7 30.4 54.1 3.60 1.36 2.45 4.07 3.16 0.73 2.56 1.78 0.63 2.64 23.0 26.7 49.7 3.61 1.72 2.81 6.06 4.76 1.51 3.30 2.75 0.80 4.05 31.4 31.7 63.1 DM basis a TEAA = total essential amino acids. TNEAA = total nonessential amino acids. TAA = total amino acids. b c 2419 Alternative protein sources for dogs Table 6. Protein quality evaluation of chicken and blood protein sources and enzyme-hydrolyzed fish protein concentrate, soybean meal, and spray-dried pork livera Item b CBM PBM SDCC SDCCL SDE PRBC SDP SDWBB EHFPC SBM SDPL PSI, protein %c IDEA valued Lysine digestibility, %e 37.5 26.6 44.4 24.2 81.2 23.9 92.9 92.5 53.4 66.8 26.4 0.54 0.43 0.64 0.47 NDf 0.54 0.58 0.45 ND 0.79 0.48 84.8 68.3 92.5 76.1 ND 84.4 88.4 72.2 ND 88.6 77.4 a Samples were analyzed in duplicate, but multiples of the same type of sample, which would allow for statistical analysis, were not analyzed. b CBM = chicken by-product meal; PBM = poultry by-product meal; SDCC = spray-dried cooked chicken; SDCCL = spray-dried cooked chicken liver; SDE = spray-dried egg; PRBC = processed red blood cells; SDP = spray-dried plasma; SDWBB = spray-dried whole beef blood; EHFPC = enzyme-hydrolyzed fish protein concentrate; SBM = soybean meal; SDPL = spray-dried pork liver. c Protein solubility index of protein in sample based on potassium hydroxide assay. d IDEA = immobilized digestive enzyme assay value used to calculate lysine digestibility. e Lysine digestibility value based on IDEA value. f ND = no data; IDEA assay not developed for this protein source. to pigs (Hansen et al., 1993), and this would affect the inclusion level allowed in a diet. Enzyme-hydrolyzed fish protein concentrate had concentrations of TAA (Table 5) lower than the CP concentration, indicating the presence of N-containing compounds that are not true protein. Fish proteins are known to be highly palatable to dogs (Heinicke, 2003). Soybean meal had higher TNEAA concentrations than TEAA concentrations. The concentration of TAA is lower than the CP concentration. The CP value for SBM used in this study was slightly less than that reported by Clapper et al. (2001) of 56.6%; however, similar TEAA concentrations were found in both studies. Soybean protein is typically added to pet foods as a complementary protein in primarily grain-based diets (Hill, 2003). Spray-dried pork liver had higher TNEAA concentrations than TEAA concentrations. Again, the concentration of TAA was lower than the CP concentration. Liver improves palatability when incorporated as 3% of the total diet or 5% of the total protein in a diet (Heinicke, 2003). The addition of liver to a pet food above a concentration of 10% needs to be closely monitored as the resulting diet may contain excessive vitamin A concentrations (Heinicke, 2003). Protein quality evaluations of the 11 protein sources studied are reported in Table 6. Protein solubility index values differed among chicken protein sources, blood protein sources, enzyme-hydrolyzed fish protein concentrate, and spray-dried pork liver. There also was large variation in PSI values within chicken protein and Table 7. Chick protein efficiency ratio (PER) assay results Treatmenta CBM PBM SDCC SDCCL SDE PRBC SDP SDWBB EHFPC SBM SDPL SEMb LSD Weight gain, g/(chickⴢd) Feed intake, g/(chickⴢd) Protein intake, g/(chickⴢd) PER 6.8 5.2 8.6 12.3 14.0 −1.3 1.1 −1.4 4.6 9.2 10.4 3.4 9.7 19.9 19.3 21.3 27.3 26.3 6.0 10.1 5.0 16.1 24.0 25.9 7.5 21.8 2.0 1.9 2.1 2.7 2.6 0.6 1.0 0.5 1.6 2.4 2.6 0.7 2.1 3.42 2.73 4.06 4.50 5.32 −2.25 1.15 −2.99 2.83 3.84 4.00 0.17 0.49 a CBM = chicken by-product meal; PBM = poultry by-product meal; SDCC = spray-dried cooked chicken; SDCCL = spray-dried cooked chicken liver; SDE = spray-dried egg product; PRBC = processed red blood cells; SDP = spray-dried plasma; SDWBB = spray-dried whole beef blood; EHFPC = enzyme-hydrolyzed fish protein concentrate; SBM = soybean meal; SDPL = spray-dried pork liver. b Pooled SEM, n = 10 chicks per treatment. blood protein sources. Spray-dried plasma and spraydried whole beef blood had PSI values higher than spray-dried egg. Egg is very close to being an ideal protein as it has a 95% biological value (FAO, 1970); therefore, it is a standard for comparison among protein sources. Enzyme-hydrolyzed fish protein concentrate, processed red blood cells, SBM, and spray-dried pork liver had lower PSI values than did spray-dried egg. The IDEA lysine digestibilities varied among chicken protein sources, blood protein sources, SBM, and spraydried pork liver, with the largest variation occurring for the chicken protein sources. The IDEA lysine digestibility was similar for spray-dried cooked chicken liver and spray-dried pork liver (average = 76.8%), for chicken-by-product meal and processed red blood cells (average = 84.6%), and for SBM and spray-dried plasma (average = 88.5%). Thus, in the protein quality evaluations there was not a clear picture defined as to the best-quality protein source. Further research into the appropriate in vitro method for assaying the protein quality of these products would benefit animal nutrition. The PER experiment (Table 7) indicated that chicken protein sources are high-quality proteins as the PER values were all greater than 2.7. Casein is considered to be a standard reference protein with a PER value of 2.5 (Munro and Allison, 1969). The highest PER value was for spray-dried egg, comparable to results of Johnson and Parsons (1997), who reported a PER value for this ingredient of 4.65. Spray-dried cooked chicken and spray-dried cooked chicken liver were not different in PER. Chicken by-product meal had a higher (P < 0.05) PER value than poultry by-product meal, which was likely the result of chicken-by product meal being ground, clean, rendered carcass parts of chicken with 2420 Dust et al. only trace amounts of feathers and blood, whereas poultry by-product meal is ground, clean, rendered carcass parts of poultry, but it includes heads, feet, viscera, and trace amounts of feathers and blood (AAFCO, 2003). The varying concentrations of poultry parts, especially feathers, in poultry by-product meal have been reported to have a negative effect on nutrient digestibilities (Morris and Balloun, 1973; Latshaw, 1990). Johnson and Parsons (1997) found poultry by-product meal had a PER value ranging from 2.43 to 2.50, depending on ash content. Feeding the blood protein sources resulted in lower PER values than the chicken protein sources. Of the three sources, spray-dried plasma had the highest PER value. Processed red blood cells and SWBB resulted in negative PER values, likely due to low feed (and protein) intakes. These diets obviously were unpalatable to the chicks (Waibel et al., 1977), and other reports have shown blood products to be unpalatable to pigs (Kratzer and Green, 1957; Pond and Maner, 1984). The blood products had low isoleucine concentrations; therefore, the diets may have been deficient in isoleucine, limiting the growth potential of the chicks. Blood products typically have a favorable profile but have low concentrations of methionine and isoleucine (Quigley et al., 2000). Practically speaking, this is of little importance as blood products are added to diets at low concentrations normally, mainly for their immune-enhancing properties for dogs and pigs (C. M. Grieshop, unpublished data, Univ. of Illinois at Urbana-Champaign; Coffey and Cromwell, 2001). Nutrient digestibilities may be affected by spray-dried plasma supplementation as Dust et al. (2003) reported that dogs fed diets supplemented with 0.5 or 1% spray-dried plasma had increased (P < 0.05) ileal digestibilities of DM, OM, CP, and acid-hydrolyzed fat; however, spray-dried plasma supplementation did not affect total-tract nutrient digestibilities. Of the other protein sources studied, spray-dried pork liver resulted in the greatest PER value, followed by SBM and enzyme-hydrolyzed fish protein concentrate. The PER value for SBM corroborates the fact that soybean protein inclusion in dog diets results in excellent CP digestibilities at the ileum (79.2 and 85.3%; Bednar et al., 2000; Clapper et al., 2001, respectively). The composition of the processed red blood cells used in palatability and digestibility studies are presented in Table 8. Processed red blood cells replaced the corn gluten meal component of the control diet such that the treatment diet contained 3% supplemental processed red blood cells. Diets had similar concentrations of DM, OM, and GE. For the processed red blood cells-containing diet, CP concentration was 6% greater and acidhydrolyzed fat content 6.5% greater than that of the control diet. Palatability results are presented in Table 9. Dogs consumed more of the control diet than the diet containing 3% processed red blood cells. According to Griffen (2003), an animal’s appetite can skew food pref- Table 8. Chemical composition of processed red blood cells (PRBC) and dietary treatments containing 0 or 3% PRBC for palatability and digestibility experiments with dogs Dietary treatment Item PRBC 0% PRBC 3% PRBC DM, % 96.1 91.2 91.4 OM, % CP, % Acid-hydrolyzed fat, % GE, kcal/g 90.8 97.2 0.9 5.4 DM basis 94.6 24.9 8.7 4.7 94.0 26.5 9.3 4.8 erence; therefore, consumption alone should not be the deciding factor regarding food preference. If the dog is hungry, it could eat equal amounts of both diets. In this experiment, there was little difference between first approach of the 3% processed red blood cells diet (51%) and the 0% processed red blood cells diet (49%). The dogs first choice for consumption was the control diet (72%) rather than the 3% processed red blood cells diet (28%). First approach and first consumed reponses are very subjective and therefore are not the best indicators of palatability. In addition, first approach and first consumption data often are difficult to measure and the repeatability of these measures is questionable (Griffen, 2003). The intake ratio for the 3% processed red blood cells treatment was 0.34. An intake ratio of greater than 0.50 implies a preference for a particular diet. Determination of the corrected intake ratio allowed for statistical evaluation of diet preference if different from zero. Corrected intake ratio data support the intake ratio data as the dogs preferred the control diet. Intake ratios are the best indicators of overall palatability preference (Trivedi et al., 2000). Based on this intake ratio and the measures discussed above, it can be concluded that the 3% processed red blood cells diet was not preferred Table 9. Palatability results obtained when feeding processed red blood cells (PRBC) to dogs (n = 20) Dietary treatment Item 0% PRBC 3% PRBC SEM P-value Amount consumed (as-fed basis), g/d Intake ratioa Corrected intake ratiob 146 0.66 0.16 72 0.34 −0.16 12.8 0.06 0.06 0.01 0.01 0.01 a Intake ratio = g of 0 or 3% PRBC-containing diet consumed/g of both diets consumed. b Corrected intake ratio = Intake ratio − 0.5, to indicate a diet preference if there was a diet preference significantly different from zero. 2421 Alternative protein sources for dogs Table 10. Nutrient intakes, apparent total tract digestibilities, and fecal characteristics of dogs fed either 0 or 3% processed red blood cell (PRBC)-containing dietsa Dietary treatment Item Daily nutrient intake DM, g 0% PRBC 3% PRBC SEMb P-valuec 285.5 286.1 21.5 0.97 289.6 27.3 75.0 1,345.7 288.0 28.1 76.8 1,349.0 21.8 2.1 5.7 101.5 0.93 0.67 0.71 0.97 83.2 86.4 90.8 83.0 85.4 82.2 85.4 91.1 80.5 84.4 0.6 0.6 0.3 0.7 0.6 0.22 0.12 0.27 0.01 0.11 182 2.8 195 3.0 15.5 0.1 0.41 0.12 DM basis OM, g Acid-hydrolyzed fat, g CP, g GE, kcal/kg Total-tract digestibility, % DM OM Acid-hydrolyzed fat CP GE Fecal characteristics Fecal output, g/d (as-is basis) Fecal scorec a Values are means, n = 14 per treatment. SEM = pooled standard error of the mean. Fecal samples were scored according to the following system: 1 = hard, dry pellets – small, hard mass; 2 = hard, formed, dry stool – remains firm and soft; 3 = soft, formed moist – softer stool that retains shape; 4 = soft, unformed – stool assumes shape of container; 5 = watery – liquid that can be poured. b c when dogs were offered a choice of a diet containing no processed red blood cells. Palatability results can be influenced by several factors, especially flavor, food texture, and size and shape of kibble (Trivedi et al., 2000). An animal that is accustomed to a certain flavor will prefer to consume a diet of similar flavor. The processed red blood cells could have provided a taste that the dogs did not prefer, thereby lowering the intake ratio. Hansen et al. (1993) reported that pigs did not find spray-dried blood meal to be palatable when initially enountered. In our study, the processed red blood cells had been processed to remove the color and odor of spray-dried blood meal, but there must have been other components that led the dogs to consume more of the 0% processed red blood cells diet than the 3% processed red blood cells diet. Nutrient intake, apparent total-tract digestibility, and fecal characteristic data are presented in Table 10. Intakes were determined during the fecal collection period. The dogs fed the 0 or 3% processed red blood cells diets consumed similar quantities of nutrients daily. Total-tract digestibilities of DM, OM, acid-hydrolyzed fat, and GE did not differ between treatments. Crude protein digestibility was slightly greater (P = 0.01) for dogs consuming the control diet than for dogs consuming the 3% processed red blood cells diet. Totaltract CP digestibility values are affected by endogenous secretions (sloughing of intestinal mucosa, endogenous proteins, intestinal bacteria, and spent enzymes) and by ingredient and/or diet processing. The results of the IDEA and PSI values indicated that processed red blood cells are a high-quality protein; however, the processed red blood cells were incorporated into the diet before extrusion and the protein quality could have been compromised by the conditions of the extrusion process. In addition, Waibel et al. (1977) suggested that decreased N digestibility may be due to cross linkages other than lysine-isopeptides. Fecal wet weight (as-is basis) and fecal score data are presented in Table 10. There were no differences in these response criteria between treatments. Fecal scores of 3 are considered ideal. Dogs in this study had close to ideal fecal scores, indicating that feeding a diet containing processed red blood cells does not alter fecal characteristics in a negative manner. Alternative protein sources can serve many functions, including provision of essential and nonessential AA to the animal, as palatants, or as immune enhancers. Chicken products, particularly eggs, are a good source of protein and also are highly palatable to dogs (Heinicke, 2003). Blood products have been shown to provide immunological enhancement, particularly use of spray-dried plasma in weanling pig (Coffey and Cromwell, 2001) and senior dog diets (our unpublished data). The immunoglobulin fraction in spray-dried plasma allows the pig to fight against pathogens (Coffey and Cromwell, 2001). Alternative protein sources are also good palatants for dog diets, particularly fish and liver protein sources (Heinicke, 2003). Compositional protein quality, palatability, and digestibility data are needed to make optimal use of the many traditional and alternative protein sources available to the pet food industry. Results of the current study indicate that based on chemical composition and protein quality data, alternative protein sources differ greatly, even within category of ingredients. The variation in protein quality assessment using the PSI, IDEA, and PER assays indicates that different fractions of protein quality may be able to be evaluated. Results of this study indicate that chicken, enzyme-hydrolyzed fish protein concentrate, spray-dried pork liver, and soybean meal protein sources have high protein quality based on the PSI, IDEA, and PER data. Nonetheless, the blood protein sources are high-quality proteins based on PSI and IDEA data but, according to the PER data, have low protein quality. In addition, blood sources suffer from poor palatability, even at low-level dietary additions. Literature Cited AACC. 1983. Approved Methods. 8th ed. Am. Assoc. Cereal Chem., St. Paul, MN. AAFCO. 2003. Association of American Feed Control Officials: Official Publication. The Association, Atlanta, GA. AOAC. 1985. Official Methods of Analysis. 14th ed. Assoc. Off. Anal. Chem., Washington, DC. AOAC. 1995. Official Methods of Analysis. 15th ed. Assoc. Off. Anal. Chem., Arlington, VA. 2422 Dust et al. Araba, M., and N. M. Dale. 1990. Evaluation of protein solubility as an indicator of over processing soybean meal. Poult. Sci. 69:76–83. Bednar, G. E., S. M. Murray, A. R. Patil, E. A. Flickinger, N. R. Merchen, and G. C. Fahey, Jr. 2000. Selected animal and plant protein sources affect nutrient digestibility and fecal characteristics of ileally cannulated dogs. Arch. Anim. Nutr. 53:127–140. Budde, E. F. 1952. The determination of fat in baked biscuit type of dog foods. J. Assoc. Off. Agric. Chem. 35:799–805. Clapper, G. M., C. M. Grieshop, N. R. Merchen, J. C. Russett, J. L. Brent, Jr., and G. C. Fahey, Jr. 2001. Ileal and total tract nutrient digestibilities and fecal characteristics of dogs as affected by soybean protein inclusion in dry, extruded diets. J. Anim. Sci. 79:1523–1532. Coffey, R. D., and G. L. Cromwell. 2001. Use of spray-dried animal plasma in diets for weanling pigs. Pig News Inf. 22:39N–47N. DeRouchey, J. M., M. D. Tokach, J. L. Nelssen, R. D. Goodband, S. S. Dritz, J. C. Woodworth, and B. W. James. 2002. Comparison of spray-dried blood meal and blood cells in diets for nursery pigs. J. Anim. Sci. 80:2879–2886. Dust, J. M., G. C. Liu, C. M. Grieshop, N. R. Merchen, J. D. Quigley, and G. C. Fahey, Jr. 2003. Effects of supplemental spray dried plasma on food intake, nutrient digestibility, and gastrointestinal microflora in healthy adult dogs. J. Anim. Sci. 86 (Suppl. 1):260. (Abstr.) FAO. 1970. The amino acid content of foods and biological data on proteins. Nutritional study # 24. Unipub, Inc., Lanham, MD. Griffen, R. 2003. Palatability testing methods: Parameters and analyses that influence test conditions. Pages 187–193 in Petfood Technology. 1st ed. J. L. Kvamme and T. D. Phillips, ed. Watt Publishing Co., Mt. Morris, IL. Hansen, J. A., J. L. Nelssen, R. D. Goodband, and T. L. Weeden. 1993. Evaluation of animal protein supplements in diets of earlyweaned pigs. J. Anim. Sci. 71:1853–1862. Heinicke, H. R. 2003. Factors affecting the palatability of canned and semi-moist petfoods. Pages 183–186 in Petfood Technology. 1st ed. J. L. Kvamme and T. D. Phillips, ed. Watt Publishing Co., Mt. Morris, IL. Hill, D. A. 2003. Fiber, texturized protein and extrusion. Pages 361– 363 in Petfood Technology. 1st ed. J. L. Kvamme and T. D. Phillips, ed. Watt Publishing Co., Mt. Morris, IL. Hutton, J. 2002. Testing of petfood preference. Pages 19–25 in Proc. Petfood Forum: Focus on Quality. Watt Publishing Co., Mt. Morris, IL. Johnson, M. L., and C. M. Parsons. 1997. Effects of raw material source, ash content, and assay length on protein efficiency ratio and net protein ratio values for animal protein meals. Poult. Sci. 76:1722–1727. Johnston, J., and C. N. Coon. 1979. The use of varying levels of pepsin for pepsin digestion studies with animal proteins. Poult. Sci. 58:1271–1273. Kats, L. J., J. L. Nelssen, M. D. Tokach, R. D. Goodband, T. L. Weeden, S. S. Dritz, J. A. Hansen, and K. G. Friesen. 1994. The effects of spray-dried blood meal on growth performance of the earlyweaned pig. J. Anim. Sci. 72:2860–2869. Kerr, C. A., R. D. Goodband, J. W. Smith, II, R. E. Musser, J. R. Bergström, W. B. Nessmith, Jr., M. D. Tokach, and J. L. Nelssen. 1998. Evaluation of potato proteins on the growth performance of early-weaned pigs. J. Anim. Sci. 76:3024–3033. Kratzer, F. H., and N. Green. 1957. The availability of lysine in blood meal for chicks and poults. Poult. Sci. 36:562–565. Latshaw, J. D. 1990. Quality of feather meal as affected by feather processing conditions. Poult. Sci. 69:953-958. Morris, W. C., and S. L. Balloun. 1973. Evaluation of five differently processed feather meals by nitrogen retention, net protein values, xanthine dehydrogenase activity and chemical analysis. Poult. Sci. 52:1075–1084. Munro, H. N., and J. B. Allison. 1969. Mammalian Protein Metabolism. Vol. 3. Academic Press, New York, NY. Murray, S. M., A. R. Patil, G. C. Fahey, Jr., N. R. Merchen, and D. M. Hughes. 1997. Raw and rendered animal by-products as ingredients in dog diets. J. Anim. Sci. 75:2497–2505. NRC. 1985. Nutrient Requirements of Dogs. Natl. Acad. Press, Washington, DC. Pond, W. G., and J. H. Maner. 1984. Swine Production and Nutrition. The AVI Publishing Co., Westport, CT. Quigley, J. D., J. M. Campbell, J. Polo, and L. E. Russell. 2004. Effects of spray-dried animal plasma on intake and apparent digestibility in dogs. J. Anim. Sci. 82:1685–1692. Quigley, J. D., C. A. Jaynes, M. L. Miller, E. Schanus, H. ChesterJones, G. D. Marx, and D. M. Allen. 2000. Effects of hydrolyzed spray dried red blood cells in milk replacer on calf intake, body weight gain, and efficiency. J. Dairy Sci. 83:788–794. Schasteen, C., J. Wu, M. Schulz, and C. M. Parsons. 2002. An enzymebased digestibility assay for poultry diets. Proc. Multi-State Poult. Feeding and Nutr. Conf. Available: http://ag.ansc.purdue.edu/poultry/multistate/MultistateIDEA2002CS.pdf. Accessed Jan. 20, 2004. Trivedi, N., J. Hutton, and L. Boone. 2000. Useable data: How to translate the results derived from palatability testing. Petfood Ind. 42:42–44. Waibel, P. E., M. Cuperlovic, R. F. Hurrell, and K. J. Carpenter. 1977. Processing damage to lysine and other amino acids in the manufacture of blood meal. J. Agric. Food Chem. 25:171–175.