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Animal Science Journal (2015) 86, 772–781

doi: 10.1111/asj.12359

ORIGINAL ARTICLE Comparison of spray-dried egg and albumen powder with conventional animal protein sources as feed ingredients in diets fed to weaned pigs Sai ZHANG,1 Xiangshu PIAO,1 Xiaokang MA,1 Xiao XU,1 Zhikai ZENG,1 Qiyu TIAN1 and Yao LI2 1

State Key Laboratory of Animal Nutrition, Ministry of Agriculture Feed Industry Centre, China Agricultural University, Beijing and 2Shendi Company, Jingmen, China

ABSTRACT We evaluated the apparent (AID) and standardized ileal digestibility (SID) of amino acids (AA) in spray-dried egg (SPE) and albumen powder (AP) compared with spray-dried porcine plasma (SDPP), dried porcine solubles (DPS) and fish meal (FM). Additionally, the effects of these egg byproducts as a replacement for conventional animal proteins on the performance and nutrient digestibility of piglets were studied. In Exp. 1, six barrows fitted with ileal T-cannulas were allotted to a 6 × 6 Latin Square design and fed six diets. The AID and SID of AA were generally higher in AP and FM (P < 0.01) than in the other protein sources. In Exp. 2, 150 piglets weaned at 21 days, were fed diets containing the five protein sources for 3 weeks. Weight gain of piglets fed SDPP was the highest among the treatments. Dry matter and protein digestibility for pigs offered SDPP were higher (P < 0.01) than those offered FM and DPS. AP decreased (P < 0.05) Escherichia coli counts in the cecum. DPS decreased (P < 0.05) serum diamine oxidase compared with SPE. In conclusion, AP and SPE are competitive with traditional animal protein sources and can be successfully fed to piglets without compromising performance.

Key words: amino acid, digestibility, egg, performance, pig.

INTRODUCTION Animal protein sources provide nutritional advantages over plant protein sources in starter pig diets (Chiba 2001). Compared with plant proteins, animal proteins are superior sources of amino acids as well as vitamins and minerals such as vitamin B12, calcium and phosphorus (Chiba 2001). Conventional animal protein sources such as spraydried porcine plasma (SDPP), dried porcine solubles (DPS) and fish meal (FM) are expensive, in short supply and variable in quality. Large amounts of egg byproducts such as spray-dried egg (SPE) and albumen powder (AP) are produced annually and are unsuitable for human consumption (Schmidt et al. 2003). These are promising ingredients for incorporation into diets fed to weanling pigs (Schmidt et al. 2003). However, despite their desirable energy content, amino acid (AA) profile and additional bioactive substances (i.e. lysozyme), they have seldom received attention as alternative feedstuffs for use in swine production. Protein sources can be well-utilized only if diets are formulated precisely and correctly (Chiba 2001). Thus, accurate nutritional information (digestible AA) on © 2015 Japanese Society of Animal Science

protein sources is vital to make appropriate adjustments in the formulation of cost-effective diets (Chiba 2001). The apparent total tract digestibility (ATTD) of AA, does not accurately represent the AA absorbed by pigs (Sauer & Ozimek 1986). Because an overwhelming majority of amino acids are absorbed in the small intestine rather than the hindgut, in which microbial fermentation deviates the AA digestibility for pigs, apparent and standardized ileal digestibility of AA are recommended (NRC 2012). Apparent (AID) or standardized ileal digestibility (SID) of AA in FM (Kim & Easter 2001), DPS (Sulabo et al. 2013), SDPP (Schmidt et al. 2003; Heo et al. 2012), SPE (Schmidt et al. 2003; Heo et al. 2012) and AP (Schmidt et al. 2003) have been evaluated. Nevertheless, these data are not totally comparable since experimental conditions vary considerably and are influenced by many different factors. To our Correspondence: Xiangshu Piao, State Key Laboratory of Animal Nutrition, Ministry of Agriculture Feed Industry Centre, China Agricultural University, Beijing 100193, China. (Email: [email protected]) Received 1 August 2014; accepted for publication 3 October 2014.

ANIMAL PROTEIN SOURCES FOR WEANED PIGS

knowledge, published information is not available about the AID and SID of AA and piglet performance determined simultaneously for SPE, AP, SDPP, DPS and FM under the same experimental conditions. Therefore, the purpose of this study was to evaluate the AID and SID of AA in SPE and AP compared with SDPP, DPS and FM. In addition, the effects of replacing conventional animal proteins (SDPP, DPS, FM) with egg byproducts (SPE, AP) on the performance, nutrient digestibility and intestinal health of weaned piglets was assessed.

MATERIALS AND METHODS General All procedures used in these experiments were approved by the China Agriculture University Institutional Animal Care and Use Committee (Beijing, China). Spray-dried egg and albumen powder used in these experiments were supplied by the Shendi Company (Jingmen, China). Spray-dried porcine plasma was provided by the NP Protein Company (Tianjin, China). Dried porcine solubles were supplied by the ZKJM Company (Beijing, China). Fish meal was imported from Peru. The analyzed nutrient composition of the protein ingredients is presented in Table 1.

Animals and experimental design Exp. 1 was conducted to evaluate the AID and SID for crude protein (CP) and AA in SPE, AP, SDPP, DPS and FM

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using chromic oxide as an indigestible marker. Six crossbred barrows (Duroc × (Landrace × Large White)), with an average body weight (BW) of 17.83 ± 1.70 kg, were surgically fitted with a T-cannula at the distal ileum using procedures adapted from Stein et al. (1998). After 10 days of recovery, the pigs were allotted to a Latin Square involving six periods and six diets. Five diets were formulated to contain each protein ingredient as the only AA source. Because FM and SDPP have less desirable palatability compared with SPE, as a result of high salt content, they are only included for 20% of the diet. The additional energy in SPE (due its high fat content) was considered when formulating the diets. A N-free diet was also included to determine the basal endogenous losses of CP and AA. The formulation of the N-free diet was adapted from Stein et al. (2007) (Table 2). Vitamins and minerals were supplemented to meet or exceed the estimated nutrient requirements for nursery pigs, as recommended by the NRC (1998). The analyzed composition of the experimental diets is shown in Table 2. Pig BW was recorded at the beginning and at the end of each period. Feed allowance was equivalent to 4% of body weight and divided into two equal meals fed at 08.00 and 17.00 hours each day. Water was available at all times throughout the experiment. Each period consisted of a 5-day adaptation period followed by 2 days collection of ileal digesta from 08.00 to 17.00 hours. A 200 mL plastic bag was attached to the open cannula using a cable tie. Bags were removed whenever they were filled with digesta, or at least every 30 min and stored at –20°C to prevent bacterial degradation of the AA in the digesta.

Table 1 Analyzed nutrient composition (% as fed) of spray-dried egg (SPE), albumen powder (AP), spray-dried porcine plasma (SDPP), dried porcine solubles (DPS) and fish meal (FM)†

Ingredient

Dry matter, % Crude protein, % Ash, % Calcium, % Total phosphorus, % Acid hydrolyzed ether extract, % Indispensable amino acids (AA), % Arginine Histidine Isoleucine Leucine Lysine Methionine Phenylalanine Threonine Tryptophan Valine Dispensable AA, % Alanine Aspartic acid Cystine Glutamic acid Glycine Proline Serine Tyrosine

SPE

AP

SDPP

DPS

FM

96.23 42.76 3.40 0.15 0.55 35.8

93.37 73.23 4.80 0.12 0.67 7.70

91.56 75.13 12.48 0.29 1.22 0.90

93.42 51.67 14.49 0.33 0.52 8.69

92.75 67.17 15.21 3.20 2.25 9.08

2.91 0.95 1.94 3.34 2.68 0.87 2.15 1.68 0.51 2.12

4.84 1.66 3.10 5.26 8.73 0.92 3.47 2.54 0.74 3.14

4.30 2.49 2.50 7.40 6.44 0.56 4.24 4.13 1.45 4.65

3.39 1.19 2.35 4.57 2.91 0.96 2.49 1.82 0.78 2.74

3.68 2.26 2.62 4.93 5.39 1.69 2.63 2.84 0.72 3.06

1.96 4.44 0.76 6.40 1.59 1.92 2.42 1.43

2.86 7.40 0.94 11.43 2.66 3.35 3.51 2.42

3.49 7.06 2.18 10.33 2.54 3.95 4.01 3.11

2.33 5.21 0.47 7.67 2.75 2.51 1.95 1.60

3.31 6.00 0.45 8.25 4.12 2.55 2.56 1.66

†All data are the results of a chemical analysis conducted in duplicate.

Animal Science Journal (2015) 86, 772–781

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774 S. ZHANG et al.

Table 2 Ingredient and nutrient content of diets fed in Exp. 1 (% as-fed)

Diet†

Cornstarch Spray-dried egg Albumen powder Spray-dried porcine plasma Dried porcine solubles Fish meal Sucrose Soybean oil Solka floc Dicalcium phosphate Ground limestone Magnesium oxide Potassium carbonate Sodium chloride Vitamin mineral premix‡ Chromic oxide Nutrient levels, % Dry matter Crude protein Acid hydrolyzed ether extract Ash Calcium Total phosphorus Indispensable amino acids (AA), % Arginine Histidine Isoleucine Leucine Lysine Methionine Phenylalanine Threonine Tryptophan Valine Dispensable AA, % Alanine Aspartic acid Cystine Glutamic acid Glycine Proline Serine Tyrosine

SPE

AP

SDPP

DPS

FM

N free

44.82 40.00 – – – – 10.00 – 2.00 1.65 0.43 – – 0.30 0.50 0.30

56.69 – 25.00 – – – 10.00 3.00 2.00 1.88 0.33 – – 0.30 0.50 0.30

61.83 – – 20.00 – – 10.00 3.00 2.00 1.55 0.52 – – 0.30 0.50 0.30

51.87 – – – 30.00 – 10.00 3.00 2.00 1.95 0.08 – – 0.30 0.50 0.30

63.70 – – – – 20.00 10.00 3.00 2.00 0.50 – – – – 0.50 0.30

76.10 – – – – – 15.00 3.00 2.00 2.40 – 0.10 0.30 0.30 0.50 0.30

92.91 17.39 15.39 3.84 0.72 0.60

91.83 19.05 4.91 4.49 0.69 0.60

90.81 15.69 2.17 5.01 0.67 0.61

91.65 15.84 6.03 6.75 0.70 0.61

90.98 14.02 4.94 3.82 0.68 0.59

90.77 – 1.95 3.42 0.69 0.58

1.12 0.44 0.83 1.44 1.17 0.36 0.90 0.71 0.20 0.90

1.23 0.45 0.82 1.46 2.33 0.23 0.89 0.70 0.20 0.88

0.79 0.52 0.53 1.54 1.32 0.15 0.86 0.86 0.25 0.96

1.01 0.42 0.73 1.43 0.92 0.35 0.78 0.58 0.22 0.84

0.72 0.46 0.57 1.06 1.15 0.42 0.55 0.61 0.15 0.66

– – – – – – – – – –

0.76 1.85 0.35 2.86 0.67 0.70 0.97 0.47

0.73 2.00 0.31 3.31 0.73 0.88 0.92 0.48

0.80 1.46 0.52 2.15 0.54 0.86 0.83 0.52

0.78 1.64 0.23 2.43 0.87 0.87 0.62 0.36

0.80 1.26 0.16 1.75 0.79 0.57 0.54 0.06

– – – – – – – –

†SPE, spray-dried egg; AP, albumen powder; SDPP, spray-dried porcine plasma; DPS, dried porcine solubles; FM, fish meal. ‡Vitamin and mineral premix provided the following per kilogram of feed: vitamin A, 12 000 IU; vitamin D, 2500 IU; vitamin E, 30 IU; vitamin B12,12 μg; vitamin K, 3 mg; d-pantothenic acid, 15 mg; nicotinic acid, 40 mg; choline chloride, 400 mg; Mn, 40 mg; Zn, 100 mg; Fe, 90 mg; Cu, 8.8 mg; I, 0.35 mg; Se, 0.3 mg.

Exp. 2 was conducted to evaluate the performance, nutrient digestibility and gut health of weanling pigs. A total of 150 pigs (Duroc × (Landrace × Large White)) with an average BW of 7.06 ± 1.56 kg weaned at 21 days of age were used in a randomized complete block design. The five dietary treatments included diets supplemented with SPE, AP, SDPP, DPS and FM. Each treatment was fed to five replicate pens with six piglets (three barrows and three gilts) per pen. Relatively simple diets were designed to give an equal contribution of CP from the assay ingredient (animal © 2015 Japanese Society of Animal Science

CP : total CP = 18%), in order to receive greater difference in response to treatments. All other nutrients were kept consistent among diets. All diets (Table 3) met or exceeded recommendations for required nutrients (NRC 1998). Values for SID AA in the formula were derived from Exp. 1. All pigs were housed in a temperature-controlled nursery room (25–27°C) and were allowed to consume diets ad libitum for 21 days. Feed disappearance and pig body weight were recorded to determine average daily gain (ADG), average daily feed intake (ADFI) and gain : feed (G:F). Animal Science Journal (2015) 86, 772–781

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Table 3

Ingredient and nutrient content of diets fed in Exp. 2 (% as-fed)

Diet† SPE Corn, yellow Soybean meal, dehulled, 47% crude protein Spray-dried egg, 43% crude protein Albumen powder, 73% crude protein Spray-dried porcine plasma, 75% crude protein Dried porcine solubles, 52% crude protein Fish meal, 67% crude protein Glucose Soybean oil Dicalcium phosphate Limestone Salt L-lysine·HCl DL-methionine L-threonine L-tryptophan Zinc oxide Vitamin mineral premix‡ Nutrient levels, % Dry matter Crude protein Ash Calcium Total phosphorus SID lysine SID methionine + cystine SID threonine SID tryptophan Digestible energy, kcal/kg

61.75 21.32 8.00 – – – –

AP

SDPP

64.32 21.16 – 4.67 – – –

DPS

63.67 21.62 – – 4.56 – –

FM

61.55 21.27 – – – 6.84 –

63.81 21.00 – – – –

4.00 0.60 1.70 0.70 0.30 0.44 0.16 0.19 0.06 0.28 0.50

4.00 1.67 1.75 0.68 0.30 0.20 0.20 0.20 0.07 0.28 0.50

4.00 2.26 1.40 0.88 0.30 0.29 0.12 0.09 0.03 0.28 0.50

4.00 1.98 1.75 0.64 0.30 0.44 0.18 0.19 0.08 0.28 0.50

5.26 4.00 2.30 1.35 0.37 0.30 0.40 0.17 0.19 0.07 0.28 0.50

88.88 19.01 5.08 0.67 0.57 1.20 0.68 0.78 0.24 3400

88.27 19.03 5.14 0.69 0.58 1.20 0.68 0.78 0.24 3400

88.55 18.99 5.27 0.68 0.60 1.20 0.68 0.78 0.24 3400

88.51 19.05 5.45 0.73 0.59 1.20 0.68 0.78 0.24 3400

88.48 18.98 5.15 0.71 0.57 1.20 0.68 0.78 0.24 3400

†SPE, spray-dried egg; AP, albumen powder; SDPP, spray-dried porcine plasma; DPS, dried porcine solubles; FM, fish meal. SID, standardized ileal digestibility. ‡Vitamin and mineral premix provided the following per kilogram of feed: vitamin A, 12 000 IU; vitamin D, 2500 IU; vitamin E, 30 IU; vitamin B12, 12 μg; vitamin K, 3 mg; d-pantothenic acid, 15 mg; nicotinic acid, 40 mg; choline chloride, 400 mg; Mn, 40 mg; Zn, 100 mg; Fe, 90 mg; Cu, 8.8 mg; I, 0.35 mg; Se, 0.3 mg.

Chemical analysis of feed, digesta and feces Feed samples were collected at the beginning of each experiment. Digesta samples in Exp. 1 were thawed, homogenized within animal and diet, lyophilized in a vacuum-freeze dryer (Tofflon Freezing Drying Systems, Shanghai, China), ground through a 1-mm screen, and mixed thoroughly. Fecal samples in Exp. 2 were heat dried (65°C, 72 h) in an oven and ground to pass through a 1-mm sieve. Chemical analysis was conducted according to the methods of the AOAC (2007). Feed, digesta and fecal samples were analyzed in terms of dry matter (AOAC 2007, 930.15), ether extract (Thiex et al. 2003), Kjeldahl N (Thiex et al. 2002), ash (AOAC 2007, 942.05), calcium (AOAC 2007, 927.02) and phosphorus (AOAC 2007, 984.27). Gross energy was determined by an automatic adiabatic oxygen bomb calorimeter (Parr 1281 Automatic Energy Analyzer, Parr, Moline, IL, USA). The chromium content in the diets, digesta and feces was measured using an atomic absorption spectrophotometer (Z-5000; Hitachi, Tokyo, Japan) according to the procedure of Williams et al. (1962). Amino acids were assayed using ion-exchange chromatography with an automatic AA analyzer (L-8900 Automatic Amino Acid Analyzer; Hitachi, Tokyo, Japan) after Animal Science Journal (2015) 86, 772–781

hydrolyzing with 6N HCl at 110°C for 24 h, excluding methionine, cysteine and tryptophan. Cystine was determined as cysteic acid and methionine as methionine sulphone after peroxidation with performic acid and pre-column derivation using phenylisothiocyanate (L-8900Automatic Amino Acid Analyzer; Hitachi, Tokyo, Japan). Tryptophan was determined after hydrolyzing with 4 N LiOH at 110°C for 22 h using high performance liquid chromatography (Agilent 1200 Series; Aligent, Santa Clara, CA, USA) (Li et al. 2014).

Microbiology analysis At the end of Exp. 2, one barrow from each replicate was killed by a lethal injection of sodium pentobarbital to obtain intestinal digesta. A modification of the method as described by Yi et al. (2013) was used to determine the populations of Lactobacilli and Escherichia coli. The microbial enumerations for digesta are expressed as log10 colony-forming units per gram.

Serum metabolites and antibody titers On day 21 (Exp. 2), blood samples (10 mL) were collected from all pigs via jugular vein puncture into vacuum container tubes (Becton Dickinson Vacutainer Systems, Franklin Lakes, NJ, USA), and immediately centrifuged at 2000 × g for © 2015 Japanese Society of Animal Science

776 S. ZHANG et al.

10 min at 5°C to recover serum, which was immediately stored at –20°C until required for analysis. Serum levels of urea nitrogen, immunoglobulins (IgA, IgG), d-lactic acid and diamine oxidase activity, were quantified using a biological analyzer (7160 Automatic Biological Analyzer; Hitachi, Tokyo, Japan) at the Sino-UK Institute of Biological Technology (Beijing, China).

Calculations In Exp. 1, the equations used to calculate the amino acid digestibility of the animal protein samples were obtained from Stein et al. (2007).

AID = [1 − ( AA digesta AA diet )(Crdiet Crdigesta )] ⋅ 100% In this equation, AID is the apparent ileal digestibility of an AA [%], AAdigesta is the AA concentration in the ileal digesta (g/kg DM), AAdiet is the AA concentration in the diet (g/kg DM), Crdiet is the chromium concentration in the diet (g/kg DM) and Crdigesta is the chromium concentration in the ileal digesta (g/kg DM). The AID of CP was also calculated using this equation. The basal endogenous loss of each AA (IAAend, g/kg of dry matter intake (DMI)) at the distal ileum was determined based on the outflow obtained when pigs were fed a N-free diet using the equation of Stein et al. (2007):

IAA end = AA digesta (Crdiet Crdigesta ) In this equation, IAAend is the basal ileal endogenous loss of an AA (g/kg DM intake). The endogenous flow of CP was also determined using the same equation. By correcting the AID of each AA for the IAAend of each amino acid, standardized ileal digestibility values (SID) were calculated using the equation of Stein et al. (2007):

had a much higher acid hydrolyzed ether extract content (35.8%) compared with the other protein sources (FM, 9.1%; DPS, 8.7%; AP, 7.7%; SDPP, 0.9%). The concentration of lysine in AP (8.7%), SDPP (6.4%) and FM (5.4%) were much higher than in DPS (2.9%) and SPE (2.7%). FM contained higher methionine (1.7%) than the other protein sources. SDPP was the highest in both threonine (4.1%) and tryptophan (1.5%). The concentrations of the other essential AA were greater in SDPP than the other protein sources except for arginine (4.8%) and isoleucine (3.1%) where AP was the highest.

AA digestibility The AID and SID of CP and AA in the test ingredients are presented in Table 4. The AID of CP was in the sequence of AP (82.1%), FM (76.8%), SDPP (74.0%), SPE (70.0%) and DPS (59.8%). AID of CP (P < 0.01) and the most indispensable AA (P < 0.01), excluding leucine, methionine, threonine and tryptophan were greater in AP than other protein sources. No difference was observed in AID of CP between AP and FM, as well as SPE and SDPP. The SID of CP was in the sequence of AP (92.0%), FM (90.2%), SDPP (86.2%), SPE (81.0%) and DPS (71.8%). The SID of CP combined with the most essential AA, was higher in AP (P < 0.01) than in the remaining protein sources. No significant difference was observed in the SID of CP among AP, FM and SDPP.

SID = AID + ( IAA end AA diet ) ⋅100% where SID is the standardized ileal digestibility of an AA (%).

Statistical analysis All data were analyzed using SAS (1999: SAS Institute Inc., Cary, NC, USA). Data in Exp. 1 were analyzed using the Proc-Mixed procedure of SAS with each pig as the experimental unit. The statistical model for AID and SID of AA had treatment as a fixed effect and period and pig as random effects. In Exp. 2, all data were analyzed as a randomized complete block design with pen as the experimental unit using the GLM procedures of SAS. The results are expressed as least squares means and standard error of the mean (SEM). In all analyses, probability values less than 0.05 were used as the criterion for statistical significance.

RESULTS Chemical analysis of animal proteins The analyzed chemical composition of the animal protein sources are presented in Table 1. SDPP had the highest CP concentration (75.1%) among the five feedstuffs (AP, 73.2%; FM, 67.2%; DPS, 51.7%; SPE, 42.8%). Fish meal contained much more calcium and total phosphorus (3.2% and 2.3%) than the other ingredients (SDPP, 0.3% and 1.2%; DPS, 0.3% and 0.5%; AP, 0.1% and 0.7%; SPE, 0.2% and 0.6%). SPE © 2015 Japanese Society of Animal Science

Animal performance and digestibility of nutrients The effects of the animal protein sources on the performance and nutrient digestibility of weanling pigs are presented in Table 5. Over the entire experimental period, ADG, ADFI and G:F were not affected by the different protein sources. Apparent total tract digestibility of DM (85.1%) of pigs offered SDPP was higher (P < 0.01) than those offered AP (82.2%), FM (81.9%) and DPS (80.2%), while ATTD of DM did not differ among pigs fed SPE (83.9%) and SDPP. Pigs supplemented with SPE (77.1%) and SDPP (76.9%) had an ATTD of CP significantly higher (P < 0.01) than those fed FM (70.5%) and DPS (67.9%). Apparent digestibility of gross energy (GE) of pigs fed SDPP (84.1%) was higher (P < 0.05) than FM (80.2%).

Microbiology of digestive contents The effects of dietary animal protein ingredients on microbial concentrations in the cecum and colon are presented in Table 6. In the cecum, pigs fed AP had fewer (P < 0.05) E. coli. counts compared with DPS. Animal Science Journal (2015) 86, 772–781

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Table 4 Apparent (AID) and standardized ileal digestibility (SID) of crude protein (CP) and amino acids (AA) in animal protein sources† fed to piglets (Exp. 1) AID of animal protein sources (%) SPE

AP b

CP,% 70.00 Indispensable AA% Arginine 80.60c Histidine 74.60b Isoleucine 73.20c Leucine 72.40b Lysine 76.20c Methionine 65.60c Phenylalanine 70.40b Threonine 67.20b Tryptophan 65.20b Valine 69.00b Dispensable, AA% Alanine 65.20b Aspartic acid 75.00b Cystine 65.80 Glutamic acid 76.80b Glycine 64.00a Proline 57.20bc Serine 66.20b Tyrosine 77.60a

SDPP

DPS

FM

SID‡ of animal protein sources (%)

SEM P-value SPE

82.14

74.00

59.83

76.83

1.78 < 0.01

92.00a 88.14a 86.57a 85.43a 93.43a 84.29a 86.43a 78.86a 74.86a 83.57a

84.67b 84.17a 80.67b 83.83a 84.17b 78.50b 82.17a 76.50a 79.83a 81.17a

74.00d 70.33b 68.83a 70.17b 71.50d 67.50c 66.17c 61.00c 61.83b 64.83b

89.50a 87.50a 84.33ab 85.67a 88.17b 88.67a 82.33a 80.00a 77.50a 83.17a

1.24 1.45 1.27 1.22 1.27 1.44 1.26 1.33 1.41 1.51

79.14a 88.29a 76.14 87.86a 72.29a 59.14bc 81.71a 90.86a

77.83a 80.17b 76.00 76.00b 61.83a 63.17ab 76.33a 87.33a

59.50b 62.83c 67.83 67.83c 51.00b 49.83c 66.00b 65.17b

83.67a 78.33b 69.00 85.17a 72.83a 69.50a 79.17a 55.17b

1.96 1.38 2.93 1.98 3.24 2.53 1.46 3.72

a

b

c

ab

AP

SDPP

DPS

FM

SEM P-value

81.00

92.00

86.17

71.83

90.17

1.81 < 0.01

< 0.01 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01

86.60c 79.20b 77.00b 76.00b 79.60c 67.00c 76.40b 76.20c 72.60b 73.40b

97.57a 93.14a 90.29a 89.29a 95.00a 86.71ab 92.14a 88.00ab 82.57a 88.29a

93.33b 88.33a 86.67a 87.50a 87.50b 82.33b 88.17a 84.17b 85.83a 85.33a

80.83d 75.67b 73.17b 74.17b 75.67c 69.00c 73.00b 72.00c 68.83b 69.83b

99.33a 92.00a 89.83a 90.67a 91.50ab 89.83a 91.67a 90.67a 87.50a 89.33a

1.24 1.37 1.31 1.23 1.29 1.42 1.30 1.32 1.42 1.49

< 0.01 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01

< 0.01 < 0.01 0.10 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01

73.20b 79.20c 74.40 80.80b 89.00a 96.80a 72.60c 79.60ab

87.57a 92.14a 85.86 91.43a 95.14a 96.43a 88.43ab 92.57a

85.33a 85.33b 81.50 81.00b 94.00a 93.67a 84.17b 88.50a

67.50b 67.50d 80.83 72.00c 70.00b 62.00b 76.00c 67.50b

91.50a 84.67b 87.00 91.17a 94.00a 94.33a 90.83a 67.33b

1.99 1.36 2.90 1.99 2.26 2.86 1.43 3.75

< 0.01 < 0.01 0.09 < 0.01 < 0.01 < 0.01 < 0.01 < 0.01

b

a

ab

c

a

†SPE, spray-dried egg; AP, albumen powder; SDPP, spray-dried porcine plasma; DPS, dried porcine solubles; FM, fish meal. ‡Values for SID were calculated by correcting the values for AID for basal ileal endogenous losses. Basal ileal endogenous losses were detemined (g/kg of dry matter intake) as CP, 20.9; Arg, 0.7; His, 0.2; Ile, 0.3; Leu, 0.6; Lys, 0.4; Met, 0.1; Phe, 0.6; Thr, 0.7; Trp, 0.2; Val, 0.5; Ala, 0.7; Asp, 0.9; Cys, 0.3; Glu, 1.2; Gly, 1.8; Pro, 4.1; Ser, 0.7; and Tyr, 0.1. a–dWithin a row, different superscripts indicate a significant difference (P < 0.05).

Table 5 Effects of dietary animal proteins† on performance and apparent digestibility (%) of nutrients in weanling pigs (Exp. 2)

Performance Weight gain, g/day Feed intake, g/day Gain : feed ATTD of nutrients (%)‡ Gross energy Dry matter Crude protein

SPE

AP

SDPP

DPS

FM

SEM

P-value

337 497 0.68

346 513 0.67

357 524 0.68

333 507 0.66

345 513 0.67

6.94 12.42 0.01

0.17 0.65 0.48

0.86 0.65 1.55

0.02 < 0.01 < 0.01

83.3ab 83.92ab 77.08a

81.06ab 82.17bc 73.23ab

84.05a 85.09a 76.85a

80.61ab 80.20c 67.86b

80.24b 81.88bc 70.54b

†SPE, spray-dried egg; AP, albumen powder; SDPP, spray-dried porcine plasma; DPS, dried porcine solubles; FM, fish meal. ‡ATTD, apparent total tract digestibility. a–cWithin a row, different superscripts indicate a significant difference (P < 0.05).

Table 6 Effects of dietary animal proteins† on microbial concentrations (log10 cfu/g of digesta) in the cecum and colon (Exp. 2)

Cecum Escherichia coli Lactobacilli Colon E. coli Lactobacilli

SPE

AP

SDPP

DPS

FM

SEM

P-value

5.32ab 7.82

4.94b 7.81

5.27ab 8.05

5.63a 7.88

5.52ab 7.99

0.13 0.23

0.04 0.93

5.57 7.91

5.16 7.87

5.49 8.05

5.70 7.73

5.51 8.20

0.15 0.13

0.22 0.20

†SPE, spray-dried egg; AP, albumen powder; SDPP, spray-dried porcine plasma; DPS, dried porcine solubles; FM, fish meal. row, different superscripts indicate a significant difference (P < 0.05).

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a,b

Within a

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778 S. ZHANG et al.

Serum metabolites and antibody titers Differences in serum urea nitrogen, immunoglobulins and d-lactic acid levels were not pronounced in the current study. Serum diamine oxidase in piglets fed DPS was lower than SPE (P < 0.05) (Table 7).

DISCUSSION Chemical analysis of animal protein sources The concentration of macro-nutrients and AA profile in SPE, FM, DPS and SDPP were generally in agreement with published values (NRC 2012). In addition, the chemical composition of AP was similar to that reported by Schmidt et al. (2003). This indicates that the animal protein sources evaluated in the current study were generally representative of previously tested samples. The nutritional composition also showed that most essential AA in SPE and AP were similar with SDPP, FM and DPS except for a higher content of lysine (8.7%) in AP, greater methionine (1.7%) concentration in FM, and relatively higher content of tryptophan (1.5%) in SDPP. This suggests that AP may serve as a more preferable AA source not only for its greater total AA, but for its lysine abundance in which most cereals are deficient, since lysine is the first limiting AA in cereal-based diets (Lewis 2001). Spray-dried egg assessed in the present study contained a wellbalanced AA profile, even though its total AA was relatively lower, partly due to its high fat content. Still, SPE can be another promising protein candidate for weanling pigs.

AA digestibility To our knowledge, the present experiment is the first to report the AID and SID of AA in SPE, AP, SDPP, DPS and FM determined simultaneously under the same environmental conditions. Values for the AID and SID of AA in FM agree with previously measured results (Cervantes-Pahm & Stein 2010; NRC 2012; Sulabo et al. 2013), and the AID or SID of AA in SDPP are in

agreement with results from Schmidt et al. (2003) and the NRC (2012). Nevertheless, the AID and SID in DPS determined in this experiment were generally lower than the values reported by Sulabo et al. (2013), while the AID of cysteine calculated for DPS in this experiment is much greater than the reported values of Sulabo et al. (2013). Therefore, it is possible that the quality of DPS used in this experiment was different from that used in previous experiments, as a result of variability in mixtures of hydrolyzed intestinal mucosa combined with other ingredients, such as carriers (soybean hull). Additionally, some differences in the AID and SID of AA in SPE have been reported by Schmidt et al. (2003) and Heo et al. (2012), with higher values for the former and lower for the latter compared with current study. Differences in the method by which AID was measured may account for this discrepancy. Schmidt et al. (2003) collected ileal digesta by killing pigs rather than adopting the T-cannula technique. In addition, Schmidt et al. (2003) was focusing on the AID of AA for the formulated diet rather than for individual protein ingredients as reported by Heo et al. (2012). Thus, the AID value is more comparable between the current study and results from Heo et al. (2012) rather than Schmidt et al. (2003). Data for the AID and SID of AA in AP are limited. In the present study, values for AID and SID of essential AA in AP agree closely with those published by Schmidt et al. (2003). The reduced digestibility of CP and AA in DPS compared with values calculated for FM was also observed by Sulabo et al. (2013). These observations may be due to trypsin and chymotrypsin inhibitors (Heugten 2001) and dietary fiber (Dilger et al. 2004) in the soybean hulls serving as carriers during the processing of DPS negatively affecting the AID of CP and AA. Likewise, Heo et al. (2012) suggested that the digestibility of AA in SPE was less than that in SDPP. Compared with other animal protein sources, egg byproducts contain more trypsin inhibitors which can decrease protein digestion, leading to lower AID and SID of AA in SPE (Kato & Matsuda 1997).

Table 7 Effects of animal proteins† on N-metabolites, intestinal barrier parameters and immunoglobulin levels in weanling pigs (Exp. 2)

N-metabolites Serum urea nitrogen (mmol/L) Intestinal barrier parameters D-lactic acid (mmol/L) Diamine oxidase (U/L) Immunoglobulin levels IgG (g/L) IgM (g/L)

SPE

AP

SDPP

DPS

FM

SEM

P-value

4.69

3.67

3.31

4.55

4.29

0.48

0.25

1.33 2.12a

1.33 1.77ab

1.27 1.73ab

1.23 1.56b

1.31 1.73ab

0.03 0.11

0.14 0.02

8.56 0.90

8.02 0.89

9.44 0.89

7.98 0.79

8.19 0.91

0.36 0.06

0.06 0.58

†SPE, spray-dried egg; AP, albumen powder; SDPP, spray-dried porcine plasma; DPS, dried porcine solubles; FM, fish meal. row, different superscripts indicate a significant difference (P < 0.05).

© 2015 Japanese Society of Animal Science

a,b

Within a

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We incorporated N-free diet to determine basal endogenous losses of AA in order to determine SID values. In fact, SID of AA is preferable in diet formulation. First, AA bioavailability is well reflected by SID values by correcting basal endogenous losses of AA, which are not negligible, particularly for glycine and proline. Moreover, SID values are additive in mixtures of feed ingredients free from dietary influences, such as dietary level of AA (Stein et al. 2007). Compared with other AA within the same ingredient, the AID of glycine and proline were relatively low, but notably, the SID of these two AA was comparable with other AA (Table 4). In this regard, greater ileal endogenous AA losses of glycine and proline were postulated. Stein et al. (1999) pointed out that glycine and proline were the major components of endogenous protein (bile and mucin) and resistant to reabsorption.

Animal performance and digestibility of nutrients Piglets fed SDPP had the highest ADG, followed by FM and AP, with SPE and DPS ranking last. Numerous substitution trials have been performed (Owen et al. 1993; Schmidt et al. 2003) to evaluate the potential of egg byproducts (SPE, AP) in replacing classical protein sources (soybean meal and SDPP). These studies suggest that SPE and AP are promising alternatives to partially replace SDPP without compromising performance. Additionally, other experiments devoted to a parallel comparison between protein sources have been reported. SDPP and FM were compared under oral challenge with E. coli K88 (Bosi et al. 2004), indicating that SDPP enjoyed a better capacity for growth improvement combined with pathogen protection relative to FM. Moreover, pigs fed SDPP had a significantly higher ADG than SPE according to the results from Che et al. (2012). Vilà et al. (2010) showed positive effects of SPE with immunoglobulins and energetic fatty acid matrix in piglet performance. It has been suggested that one possible reason for the improved performance due to dietary SDPP and AP is the increased digestibility of DM and CP. In this sense, DPS demonstrated relatively decreased ADG likely due to its reduced nutrient digestibility. The fact that the ADG response to SPE and FM was not consistent with changes in apparent digestibility was probably because of the prevalence of Lactobacillus in the colon of piglets fed FM.

Microbiology of digestive contents Anderson et al. (2000) emphasized that intestinal microbes, regulated by dietary manipulation, play a pivotal role in body health. Our results demonstrated that AP can effectively restrict the prevalence of E. coli Animal Science Journal (2015) 86, 772–781

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in the cecum, followed by SDPP and SPE. The current work is not consistent with a previous study (Schmidt et al. 2003) where investigators found that AP was comparable with SDPP in inhibiting Enterobactericea. A recent in vitro study (Champagne et al. 2014) has suggested that SDPP can affect the composition of the intestinal microflora which is similar to our results. However, previous a in vivo study (Van Dijk et al. 2002), concluded that SDPP could not prevent the colonization of a pathogenic E. coli. SDPP has been regarded as an alternative for antibiotics because of its abundance of immunoglobulins (Niewold et al. 2007; Champagne et al. 2014), and lysozyme (Champagne et al. 2014), which directly or indirectly inhibit bacterial prevalence. Meanwhile, AP has been reported (Schmidt et al. 2003) to have a competitive capacity for bacteriostasis compared with SDPP, probably for its lysozyme effect. Current results for DPS manifest a relatively unfavorable microbial regulation in the piglets’ intestine. Dried porcine solubles are abundant in highquality small peptides which are known to promote performance by means of increased digestibility (Maxwell & Carter 2001) and gut health through improved intestinal morphology. However, seldom have studies reported that piglets fed DPS have altered microbial characteristics in the digestive system. Deficient in antimicrobial agents, DPS did not manifest a promising potential in this regard. In the present trial, FM enhanced the prevalence of lactobacilli in the colon. It was probably the fish oil (9.1%), reported as a microbial regulator in gut due to its n-3 fatty acid content (Shin et al. 2007), that modified the growth of lactobacilli in the current study.

Serum metabolites and antibody titers Given the improvements in growth rate of pigs fed SDPP, it is reasonable to expect alterations in serum urea nitrogen which can reflect the oxidation of dietary AA in piglets (May et al. 2012). There was no significant difference in serum urea nitrogen among the treatments. However, piglets offered SDPP had a relatively lower serum urea nitrogen level, indicating more efficient nitrogen utilization and better performance (Coma et al. 1995). Diamine oxidase, an enzyme synthesized in the intestinal mucosa of mammalian species, is a marker for monitoring mucosal injury and maturation. The damaged mucosa release diamine oxidase into the blood, indicating dysfunction in the intestinal barrier (Luk et al. 1980). Unlike diamine oxidase which is generated in the intestinal mucosa, d-lactic acid is mainly produced by intestinal microbes. A rise in d-lactic acid detected in serum has been regarded as a sign for loss of mucosal integrity (Vella & Farrugia 1998). In the current study, piglets fed DPS diets had a lower serum diamine oxidase level than the other © 2015 Japanese Society of Animal Science

780 S. ZHANG et al.

dietary treatments, indicating the intestinal barrier was well-maintained in this treatment. This is probably because of the bioactive substances which exist in DPS that hasten the maturation of the gut (Shurson & Johnston 1998). In contrast, piglets offered the SPE diet demonstrated the highest diamine oxidase level in the serum. The mechanism for this result is not known but egg allergy may serve as a possible explanation (Escribano et al. 1998). The present results show that piglets fed SDPP had a higher serum IgG than the remaining treatments. It is expected that immunoglobulins in SDPP can optimize performance through passive immunity (Coffey & Cromwell 1995), which closely agrees with our results. Egg proteins may not provide the same immunological benefits to the pig that SDPP provides because the hen encounters different diseases than the pig (Shurson & Johnston 1998). This idea coincides with the serum IgG content between piglets offered SDPP and SPE from the current study.

Conclusion Collectively, AP and SDPP are the most competitive protein ingredients among the tested protein sources in terms of performance due to their high digestible nutrient content, better AA utilization and abundance of bioactive substances (i.e. immunoglobulins, lysozyme) which regulate immune levels and intestinal microflora. SPE is high in digestible energy and protein and is comparable to FM for its desirable SID of AA. Thus, AP and SPE are competitive with traditional animal protein sources and can be successfully fed to weaned piglets without compromising performance.

ACKNOWLEDGMENTS The investigation was financially supported by the National Natural Science Foundation of China (No. 31372316).

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