METABOLISM AND NUTRITION

Effect of four processed animal proteins in the diet on digestibility

and performance in laying hens

M. M. van Krimpen,*1 T. Veldkamp,* G. P. Binnendijk,* and R. de Veer†

*Wageningen UR Livestock Research, PO Box 65, NL-8200 AB Lelystad, the Netherlands;

and †Animal Nutrition Group, Department of Animal Sciences, Wageningen University,
PO Box 338, NL-6700 AH Wageningen, the Netherlands

ABSTRACT An experiment was performed to investi-
gate the effect of animal vs. vegetable protein sources
in the diet of laying hens on the development of hen
performance. A diet containing protein sources of only
vegetable origin was compared with 4 diets, each con-
taining 1 of 4 processed animal proteins (PAP). Two
ied from 15.4 (Daka-40) to 28.3 g/kg (Sonac-50). Sub-
sequently, the effect of a control diet (without PAP)
vs. 4 PAP diets (50 g/kg of CP of animal origin from
the same batches as used in the digestibility study) on
performance was determined. All diets were isocaloric
(AMEn = 2,825 kcal/kg) and isonitrogenous (digestible

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PAP (Daka-58 and Sonac-60) were classified as meat
meals, and the remaining 2 (Daka-40 and Sonac-50)
were classified as meat and bone meals. First, fecal di-
gestibility of nutrients in the PAP was determined in
Lohmann Brown layers. Hens (n = 132) were housed in
22 cages (6 hens/cage) and allotted to 5 dietary treat-
ments. In the PAP diets (4 replicates/treatment), 100
g/kg of CP of animal origin was added, replacing soy-
bean meal and corn (Zea mays) in the basal diet (6
replicates/treatment). The PAP sources differed largely
in chemical composition and digestibility coefficients.
Energy content (AMEn) varied from 1,817 (Daka-40)
to 3,107 kcal/kg (Sonac-60), and digestible lysine var-
Key words: digestibility, laying hen, performance, processed animal protein
lysine = 6.8 g/kg). Hens were housed in 40 floor pens
(12 hens/pen, 8 pens/treatment) from 20 to 40 wk of
age. Feed intake levels of the hens fed the meat and
bone meal diets were reduced compared with those of
hens fed the meat meal diets, whereas the feed intake
level of hens fed the control diet was intermediate. Lay-
ing hen performance differed between treatments, be-
ing was most favorable for the Sonac-50 treatment and
most adverse for the Daka-40 treatment. Differences in
laying hen performance seemed to be related partly to
differences in feed intake and corresponding amino acid
intake.
2010 Poultry Science 89:2608–2616
doi:10.3382/ps.2010-00696

INTRODUCTION nonruminants (Haugen and Pettigrew, 1985). After the

Nutritionists have used processed animal proteins
(PAP) in poultry diets for many years (Kratzer and
Davis, 1959; Skurray, 1974; Waldroup and Adams,
1994; Dale, 1997; Parsons et al., 1997). The primary
advantages associated with the use of PAP in poultry
diets have been the high digestibility of amino acids, as
well as the bioavailability of phosphorus in PAP (Wal-
droup and Adams, 1994; Sell and Jeffrey, 1996; Parsons
et al., 1997). By 1950, identification of vitamin B12 as
the animal protein factor, and its commercial synthe-
sis, made it possible to develop diets without PAP for
©2010 Poultry Science Association Inc.
Received February 10, 2010.
Accepted August 29, 2010.
1 Corresponding author: marinus.vankrimpen@wur.nl
ruminant-to-ruminant ban (1989 in the Netherlands,
1994 across the European Union), inclusion of PAP in
poultry rations increased because of its favorable sup-
ply and pricing. The relatively high inclusion level of
PAP in poultry diets remained in place until the total
ban on meat and bone meals in 2001.
Since the total ban, adverse effects, such as reduced
egg production, increased susceptibility to chronic en-
teritis, and higher incidences of feather-pecking behav-
ior and cannibalism, have been reported in practice. To
date, however, it has not been demonstrated whether
these supposed effects are directly related to the chang-
es in dietary protein source. In 2005, liberalization of
the European Union regulations was announced regard-
ing the use of PAP in the diets of nonruminant ani-
mals appropriate for human consumption (C3 material,
pigs and poultry; The TSE (transmissible spongiform
encephalopathies) Roadmap; European Commission,

2608
 VEGETABLE VERSUS ANIMAL PROTEIN IN LAYER DIET
2005). The authors argued for the reuse of proteins
in nonruminant diets, provided appropriate analytical
methods are available to distinguish the origin of the
different proteins. Because of the species-to-species ban,
only meat of pork origin may be included in poultry
diets. In digestibility studies performed earlier, how-
ever, the origin of the tested PAP was rarely reported.
Therefore, the nutritional value of pork meat meal in
laying hen diets is unknown. The aim of this study was
to determine the nutritional value of 4 different PAP
sources (experiment 1) and the effects of these PAP
sources on performance (experiment 2) in laying hens.
MATERIALS AND METHODS
Materials and Methods in Experiment 1
(Digestibility Study)
2609
Experimental Design. A fecal digestibility study was
conducted in laying hens from 24 to 27 wk of age. The
experimental period began at 24 wk of age and lasted
until the end of wk 27 of age (a period of 4 wk). From
wk 26 onward, the hens received the experimental diets.
In this experiment, fecal digestibility of 4 different pork
meat meals was determined: 1) Sonac-50 (Sonac BV,
a Vion Ingredients Company, Son, the Netherlands),
2) Sonac-60 (Sonac BV), 3) Daka-40 (Daka Proteins,
Ringsted, Denmark), and 4) Daka-58 (Daka Proteins).
These PAP sources varied in chemical composition,
thereby covering the range of commercially available
pork meat meal quality, as shown in Table 1. Crude
protein content, for instance, ranged from 417 to 617 g/
kg, whereas ash content ranged from 183 to 437 g/kg.
In the preexperimental phase (from wk 17 to 23),
a commercial layer meal diet (2,825 kcal/kg, 6.7 g/kg
of digestible lysine) was fed. From wk 24 onward, the

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Housing, Birds, and Management. At 17 wk of
age, a total of 160 laying hens (ISA Brown strain) that
had passed a visual health inspection were allotted to 1
of 5 dietary treatments: 1 basal diet without PAP and
4 PAP-enriched diets. The hens were housed in battery
cages (65 × 75 cm) with 6 hens/cage in a climate-con-
trolled poultry house. The basal treatment was replicat-
ed 6 times (36 hens), whereas the PAP-enriched treat-
ments were replicated 4 times (24 hens). An additional
24 hens were kept as reserve animals until the moment
the experimental diets were provided. Temperature and
RH of the experimental room were registered continu-
ously. The target temperature was maintained at 20°C,
whereas an L:D schedule of 16 h of light and 8 h of
darkness was applied. To realize a constant feed intake
during the light period, an intermittent L:D schedule of
15 min of light and 45 min of dark was applied in this
period. All diets and water were provided ad libitum.
respective experimental meal diets were fed. The main
ingredients of the basal diet were corn (Zea mays; 747
g/kg) and heat-treated soybean meal (143 g/kg). The
PAP diets were a mixture of the basal diet and one
of the pork meat meals. Each PAP-enriched diet con-
tained approximately 100 g/kg of CP of animal origin.
Therefore, inclusion levels ranged from 164 g/kg in the
Sonac-60 and Daka-58 diets to 220 g/kg in the Daka-40
diet. Furthermore, some vitamins, minerals, and syn-
thetic amino acids were added to the diet to meet NRC
(1994) requirements. The basal diet was formulated as
a standard diet (ME 2,850 kcal/kg, digestible lysine
5.5 g/kg). Titanium oxide (1 g/kg) was included in the
diets as indigestible marker. Dietary ingredients and
calculated nutrients of the diets (in g/kg) are shown in
Table 2.
Excreta Collection and Apparent Digestibility
Calculations. In wk 27, representative samples of ex-

Table 1. Analyzed nutrients of the processed animal proteins used in experiment 1 (g/kg)1
Meat and bone meal Meat meal

Nutrient2 Daka-40 Sonac-50 Sonac-60 Daka-58

DM (g/kg) 968.7 977.5 963.9 974.8

Ash (g/kg) 437.0 336.2 182.6 251.6
Organic matter (g/kg) 531.7 641.3 781.3 723.2
CP (g/kg) 416.8 558.1 616.8 596.5
Fat (g/kg) 99.1 95.9 116.5 118.2
Gross energy (kcal/kg) 3,241 3,845 4,739 4,507
Calcium (g/kg) 160.5 116.0 52.0 82.2
Magnesium (g/kg) 3.6 2.6 2.2 2.3
Phosphorus (g/kg) 77.3 58.6 29.5 42.1
Sodium (g/kg) 7.0 6.0 6.9 7.0
Potassium (g/kg) 4.0 4.3 7.6 5.8
Amino acid        
  Lysine (g/kg) 19.7 30.9 32.5 29.4
  Methionine (g/kg) 4.9 8.5 9.3 8.2
  Cysteine (g/kg) 1.5 3.7 5.3 2.7
  Methionine + cysteine (g/kg) 6.3 12.2 14.6 10.9
  Threonine (g/kg) 11.4 17.4 20.7 18.1
  Tryptophan (g/kg) 2.1 4.0 5.1 4.2
1Sonac-50 and Sonac-60 were provided by Sonac BV (Son, the Netherlands), and Daka-40 and Daka-58 were
provided by Daka Proteins (Ringsted, Denmark).
2All samples were analyzed in duplicate, after which values were averaged.
2610 van Krimpen et al.
Table 2. Dietary ingredients and analyzed and calculated nutrients of the diets used in experiment 1 (g/kg, as-fed basis)1
Meat and bone meal Meat meal

Item Basal Daka-40 Sonac-50 Daka-58 Sonac-60

Ingredient (g/kg)          
  Corn 746.8 643.4 646.9 650.0 639.0
  Soybean meal, heat-treated 142.6 122.7 123.4 124.0 121.9
  Processed animal protein
   Sonac-50     186.5    
   Sonac-60         163.9
   Daka-40   220.0      
   Daka-58       163.5  
  Limestone 60.8   4.2 25.4 37.3
  Oyster shells 30.4 6.6 30.0 30.0 30.0
  Monocalcium phosphate 7.2        
  Salt 4.3 1.3 3.0 1.1 1.9
  Premix laying hens2 5.0 5.0 5.0 5.0 5.0
  Titanium oxide 1.0 1.0 1.0 1.0 1.0
  l-Lysine hydrochloride 0.9        
  dl-Methionine 0.9        
  l-Tryptophan 0.1        
Analyzed nutrient3 (g/kg)          

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  DM 887 896 898 896 895
  Ash 112 122 120 113 129
  CP 132 205 216 211 235
  Fat 28 46 39 48 51
  Lysine 6.58 8.94 10.93 10.22 10.48
  Methionine + cysteine 5.21 4.98 5.42 5.43 5.93
  Threonine 4.84 6.55 7.44 7.27 7.79
  Tryptophan 1.55 1.57 1.84 1.75 1.95
Calculated nutrient (g/kg)          
  Crude fiber 22 21 19 21 19
  Starch 457 394 397 398 392
  Sugar 22 19 19 19 19
  Calcium 36 40 36 36 36
  Phosphorus 4.6 18.1 15.8 9.1 5.7
  Available phosphorus 2.3 10.2 8.8 4.8 2.3
1Sonac-50 and Sonac-60 were provided by Sonac BV (Son, the Netherlands), and Daka-40 and Daka-58 were provided by Daka Proteins (Ringsted,
Denmark).
2Provided the following nutrients per kilogram of diet: vitamin A, 12,000 IU; vitamin D , 2,400 IU; vitamin E, 40 mg; vitamin B , 4.8 mg; vitamin
3 1
B2, 12 mg; d-panthothenic acid, 16 mg; niacinamide, 48 mg; vitamin B6, 5.6 mg; folic acid, 1.8 mg; vitamin B12, 25 μg; vitamin C, 100 mg; biotin, 0.1
mg; vitamin K3, 4.8 mg; choline chloride, 260 mg; copper, 8 mg (as CuSO4·5H2O); iron, 65 mg (as FeSO4·7H2O); manganese, 65 mg (as MnO2); zinc,
50 mg (as ZnSO4); cobalt, 0.4 mg (as CoSO4·7H2O); iodine, 1 mg (as KI); selenium, 0.4 mg (as Na2SeO3·5H2O).
3All samples were analyzed in duplicate, after which values were averaged.

creta were collected semiquantitatively per cage over a
period of 4 × 24 h to determine AMEn and apparent
digestibility of DM, ash, fat, and amino acids. Excreta
samples were frozen and stored at −18°C until chemical
analysis. The AMEn content was determined by cor-
recting AME values to zero nitrogen values (Hill and
Anderson, 1958). Therefore, nitrogen retention (nitro-
gen intake minus nitrogen excretion; De Jonge et al.,
2000) was multiplied by the enthalpy of oxidation of
uric acid (8.7 kcal/g) and subtracted from AME. Ap-
parent digestibility of nutrients was calculated using
the following equation:
AD (%) = {1 – [(Tifd/Tifc × Nfc/Nfd)]} × 100, [1]
where AD is the apparent digestibility of nutrients,
Tifd and Tifc represent titanium in feed and excreta,
respectively, and Nfd and Nfc are the nutrients in feed
and excreta, respectively. The AMEn and digestibility
coefficients of the PAP sources were calculated from
the differences in AMEn and digestibility coefficients
between the basal and PAP-enriched diets.
Analytical Procedures. After defrosting, excreta
samples were homogenized and subsequently, represen-
tative samples were taken. All samples were analyzed
in duplicate. For determination of the DM content in
feed and excreta, samples were freeze-dried accord-
ing to International Organization for Standardization
(ISO) method number 6496 (International Organiza-
tion for Standardization, 1998b). After freeze-drying,
samples were ground to pass a 1-mm screen and kept
for analysis. Air-dried samples were dried in a forced-
air oven at 103°C to a constant weight according to
ISO method number 6496 (International Organization
for Standardization, 1998b). Kjeldahl nitrogen content
in feed and excreta was measured according to ISO
method number 5983 (International Organization for
Standardization, 1997) in fresh samples. Crude protein
content was calculated as nitrogen × 6.25. Fecal nitro-
gen content was determined according to the method
 VEGETABLE VERSUS ANIMAL PROTEIN IN LAYER DIET
of De Jonge et al. (2000). Amino acids in feed and
excreta (except tryptophan) were determined after hy-
drolysis with 6 M hydrochloric acid during 23 h. The
hydrolysate was adjusted to pH 2.2. Before hydrolysis,
sulfur-containing amino acids were oxidated overnight
with performic acid/phenol at 0°C and neutralized with
sodium disulfite. Amino acids were separated by ion-
exchange chromatography and detected photometri-
cally after postcolumn derivatization with ninhydrin by
using an automated amino acid analyzer at 570 nm and
at 440 nm for proline according to ISO method num-
ber 13903 (International Organization for Standard-
ization, 2005a). Tryptophan was released by alkaline
hydrolysis with lithium hydroxide under nitrogen pres-
sure at 120°C for 16 h. Tryptophan was separated by
reversed-phase C18 HPLC with fluorescence detection
with acetate/methanol as eluants and identified and
quantified by UV detection at 280 nm according to ISO
method number 13904 (International Organization for
Standardization, 2005b). Crude fat content in feed and
excreta was determined after acid hydrolysis according
to ISO method number 6492 (International Organiza-
tion for Standardization, 1999). For determining crude
ash content, feed and excreta samples were incinerated
at 550°C in a muffle furnace according to ISO method
number 5984 (International Organization for Standard-
ization, 2002). Gross energy of the diet and feces was
determined using an adiabatic bomb calorimeter (IKA-
C5003, Janke & Kunkel, Staufen, Germany) accord-
ing to ISO method number 9831 (International Orga-
nization for Standardization, 1998a). Titanium oxide
was determined according to the method developed by
Short et al. (1996) and further refined by Myers et al.
(2004). This method is based on digestion of the sam-
ple in sulfuric acid and addition of hydrogen peroxide
to produce an intense orange-yellow color that is read
colorimetrically at 408 nm with a UV-visible spectro-
photometer (CARY 50 probe, Varian, Palo Alto, CA).
Materials and Methods in Experiment 2
(Performance Study)
Housing, Birds, and Management. A total of 480
non-beak-trimmed 21-wk-old layers (Isa Brown strain)
were housed in 2 climate-controlled rooms. Each room
contained 20 floor pens (0.90 × 1.50 m), and each pen
contained 12 hens. The experimental period was from
21 to 40 wk of age. The pens were built of wire and
hens could see their flock mates in other pens. Each
pen contained 4 perches, a feeding trough (length 100
cm), and 3 nipple drinkers. Sand was used as litter on
the floor. A laying nest was placed outside each pen.
Throughout the experiment, litter quality was main-
tained by adding new sand monthly. At the beginning
of the experiment, mean BW of the hens was 1,484 ±
52 g (mean ± SD). Feed and water were provided for
ad libitum consumption. Temperature was set at a con-
stant value of 21°C. Hens received an L:D schedule of
2611
16 h of light and 8 h of dark, whereas the photoperiod
lasted from 1 to 17 h. Health status of the hens was
monitored daily.
Experimental Design. Pens were allotted to 1 of 5
dietary treatments, and each treatment had 8 repli-
cates, which were equally divided over the 2 rooms. In
this study, a control diet without PAP was compared
with 4 PAP diets, thereby using the same PAP batches
as used in the digestibility experiment: Sonac-50 (treat-
ment 2), Sonac-60 (treatment 3), Daka-40 (treatment
4), and Daka-58 (treatment 5). The nutritional values
of these PAP sources (Table 1) that were determined
in the digestibility study were used to optimize the di-
ets for the performance study. All diets were isocaloric
(2,825 kcal/kg), had similar digestible essential amino
acid profiles, and met NRC (1994) requirements (Table
3). Each PAP-enriched diet contained 50 g/kg of pro-
tein of animal origin. Because of differences in protein
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content, PAP inclusion ranged from 8.1 (Sonac-60)
to 12.0% (Daka-40). Compared with the control diet,
contents of soybean meal, rapeseed meal, oyster shells,
and monocalcium phosphate were reduced in the PAP
diets, whereas content of wheat middlings increased.
Moreover, to balance the relatively low digestible lysine
content in Daka-40 and Daka-58, part of the wheat in
diets 4 and 5 was replaced by peas.
Observations. Feed consumption and hen perfor-
mance per pen were recorded weekly. All hens were
weighed in 4-wk intervals (average per pen). For the
trait total egg mass, the entire egg mass production was
calculated, assuming that shell-less and cracked eggs
had the same weight as the mean normal egg weight
of that specific pen in that week. Litter wetness was
assessed as percentage of floor area that was visually
observed as wet.
Statistical Analysis. The REML variance compo-
nent analysis procedure tested the effect of nutritional
factors on the determined traits by using model [2]:
Yij = μ + weeki + treatmentj + week
× treatment + eij, [2]
where Yij is the dependent variable, μ is the overall
mean, weeki is the fixed effect of week i (i = 21, 22, …
40), treatmentj is the fixed effect of dietary treatment
j (j = 1, 2, … 5), and eij is the error term (Genstat,
2002). The effects of room and pen were added to the
random terms of the model. The P-values for week,
treatment, and week × treatment are presented. Means
were considered significantly different when P < 0.05.
RESULTS
Results of Experiment 1
(Digestibility Study)
The digestibility coefficients of the 4 PAP sources
showed a large variation (Table 4). Dry matter digest-
2612 van Krimpen et al.

ibility ranged from 38.4 (Daka-40) to 64.3% (Sonac-60). Results of Experiment 2

Digestibility of CP ranged from 81.4 (Daka-40) to 90.1%
(Sonac-50). Digestibility of gross energy ranged from
56.0% in Daka-40 to 66.6% in Sonac-50. The Sonac-50,
Sonac-60, Daka-40, and Daka-58 had AMEn values of
2,566, 3,107, 1,819, and 2,801 kcal/kg, respectively. In
general, digestibility of amino acids was lowest in Daka-
40 and highest in Sonac-50.
(Performance Study)
Feed intake differed between treatments (P = 0.005),
being highest for the Sonac-60 treatment (124.5 g/hen
per d) and lowest for the Sonac-50 (118.9 g/hen per
d) and Daka-40 treatments (119.1 g/hen per d; Table
5). Feed intake levels of the control diet and the Daka-

Table 3. Dietary ingredients and analyzed nutrients of the diets used in experiment 2 (g/kg, as fed basis)1
Meat and bone meal Meat meal

Item Control Daka-40 Sonac-50 Daka-58 Sonac-60

Ingredient (g/kg)
  Corn 400.0 400.0 400.0 400.0 400.0
  Wheat 211.0 159.0 190.5 150.5 192.2
  Soybean meal extract (dietary fiber <50) 129.7 48.6 62.2 47.4 66.1

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  Peas (CP <220) 75.0 125.0 75.0 125.0 75.0
  Oyster shells 72.5 25.7 48.9 58.2 64.8
  Rapeseed meal extract (CP <380) 30.0 30.0 30.0 30.0 30.0
  Wheat middlings 0.0 52.3 65.5 65.8 50.9
  Processed animal protein
   Sonac-50 0.0 0.0 89.6 0.0 0.0
   Sonac-60 0.0 0.0 0.0 0.0 81.1
   Daka-40 0.0 120.0 0.0 0.0 0.0
   Daka-58 0.0 0.0 0.0 83.8 0.0
  Corn gluten meal 24.9 0.0 0.0 0.0 0.0
  Limestone 20.0 20.0 20.0 20.0 20.0
  Soybean oil 17.9 9.9 9.7 9.8 10.0
  Monocalcium phosphate 8.2 0.0 0.0 0.0 1.1
  Premix for laying hens2 5.0 5.0 5.0 5.0 5.0
  Salt 3.5 1.3 2.1 2.0 2.1
  dl-Methionine 1.2 1.9 1.4 1.7 1.5
  l-Lysine 1.0 0.6 0.0 0.4 0.1
  l-Tryptophan 0.1 0.3 0.1 0.2 0.1
  l-Threonine 0.0 0.4 0.0 0.2 0.0
Analyzed nutrient3 (g/kg)          
  DM 889 888 890 890 891
  Ash 130 120 126 127 129
  CP 159 163 167 165 168
  Organic matter 758 768 764 764 762
  Crude fat 47 48 47 47 47
  Crude fiber 36 30 31 33 33
  Dietary fiber 516 527 520 519 515
  Starch 353 381 375 365 364
  Reducing sugars4 31 25 26 25 27
  Starch + sugar 384 405 401 390 391
  Calcium 41.82 39.07 39.69 41.12 41.06
  Magnesium 1.81 1.85 1.83 1.84 1.93
  Total phosphorus 4.72 12.16 8.20 6.83 5.79
  Sodium 1.76 1.43 1.57 1.54 1.66
  Potassium 5.98 5.71 5.61 5.88 6.02
  Lysine 8.21 8.39 8.01 7.99 8.29
  Methionine 3.36 3.59 3.63 3.76 3.82
  Cysteine 2.43 2.03 2.24 2.02 2.33
  Methionine + cysteine 5.79 5.62 5.87 5.78 6.15
  Threonine 5.69 5.90 5.87 5.60 6.05
  Tryptophan 1.87 1.75 1.68 1.74 1.86
Calculated nutrient          
  AMEn (kcal/kg) 2,825 2,825 2,825 2,825 2,825
  Available phosphorus (g/kg) 2.80 6.86 4.39 2.80 2.80
1Sonac-50 and Sonac-60 were provided by Sonac BV (Son, the Netherlands), and Daka-40 and Daka-58 were provided by Daka Proteins (Ringsted,
Denmark).
2Provided the following nutrients per kilogram of diet: vitamin A, 12,000 IU; vitamin D , 2,400 IU; vitamin E, 40 mg; vitamin B , 4.8 mg; vitamin
3 1
B2, 12 mg; d-panthothenic acid, 16 mg; niacinamide, 48 mg; vitamin B6, 5.6 mg; folic acid, 1.8 mg; vitamin B12, 25 μg; vitamin C, 100 mg; biotin, 0.1
mg; vitamin K3, 4.8 mg; choline chloride, 260 mg; copper, 8 mg (as CuSO4·5H2O); iron, 65 mg (as FeSO4·7H2O); manganese, 65 mg (as MnO2); zinc,
50 mg (as ZnSO4); cobalt, 0.4 mg (as CoSO4·7H2O); iodine, 1 mg (as KI); selenium, 0.4 mg (as Na2SeO3·5H2O).
3All samples were analyzed in duplicate, after which values were averaged.
4Mono- and disaccharides as glucose units.
 VEGETABLE VERSUS ANIMAL PROTEIN IN LAYER DIET
58 diet were intermediate. Egg weight of hens fed the
Daka-58 diet (58.4 g/egg) was significantly (P = 0.014)
lower compared with the egg weight of hens fed the
control diet and the Sonac-60 diet (59.8 to 59.9 g/egg),
whereas egg weight of hens fed the Daka-40 or Sonac-50
diet was intermediate (58.7 and 59.4 g/egg, respective-
ly). Differences between treatments, however, were not
constant over time (treatment × week interaction; P
< 0.001). Egg mass levels differed between treatments
(P = 0.005) and ranged from 46.4 (Daka-40) to 49.2 g/
hen per d (Sonac-50). The amount of feed required to
produce 1 kg of eggs also differed between treatments
(P = 0.049) and ranged from 2.13 (Sonac-50) to 2.30 kg
(Sonac-60). Rate of lay and hen BW were not signifi-
cantly affected by dietary treatments. All performance
traits were significantly affected by week, although no
significant week × treatment interactions were found,
except for egg weight. The percentage of wet litter in
the pens was significantly lower for the Sonac-50- and
Daka-40-fed hens compared with hens in the other
treatments. Average litter wetness was extremely high
in wk 23 (56%), but then ranged from 19.5 (wk 28) to
25.6% (wk 30).
DISCUSSION
Nutritional Value of PAP Sources
The tested PAP sources largely varied in chemical
composition, digestibility of nutrients, and AMEn val-
ue, which is in line with earlier findings in the literature
(Hendriks et al., 2002, 2004). In particular, the ash and
protein contents of the PAP sources showed large varia-
tions. Ash content ranged from 182.6 g/kg in Sonac-60
to 437.0 g/kg in Daka-40, whereas protein content
ranged from 416.8 g/kg in Daka-40 to 616.8 g/kg in
Sonac-60. Depending on the ratio of bone to soft tissue
used in processing, the finished product was designated
as meat meal (containing >55% CP and <4.4% phos-
phorus) or meat and bone meal (containing <55% CP
and >4.4% phosphorus; Ravindran and Blair, 1993).
On the basis of this definition, Sonac-60 and Daka-58
were classified as meat meals, and Sonac-50 and Daka-
40 were classified as meat and bone meals.
The bone fraction is one of the constituents that
can affect the composition and protein quality of meat
and bone meal. Bones contain a high collagen content
of approximately 83% (Eastoe and Long, 1960). Be-
cause of their high collagen content and poor amino
acid balance, any increase in bone content in the raw
materials may have a negative effect on protein quality.
Collagen and gelatin (refined collagen) are deficient in
most essential amino acids, such as tryptophan, sulfur
amino acids, and isoleucine, whereas they oversupply
hydroxyproline, proline, and glycine (Boomgaardt and
Baker, 1972; Berdanier, 1998). The amino acid profile
of the Daka-40 tested showed that this source con-
tained a high amount of collagen. In comparison, soft
offal (muscle, gut, and stomach) contains much higher
2613
levels of essential amino acids and will produce meat
meals of greater nutritional value. An increased bone or
ash content negatively affects protein and energy con-
centrations (Dale, 1997; Mendez and Dale, 1998; Wang
and Parsons, 1998a). In line with these findings, in the
current study, a linear negative regression was found
between ash content of the PAP and the AMEn value
(P < 0.001, R2 = 84.0). The CVB (the Dutch office for
feed evaluation) developed a linear regression equation
to calculate AMEn values of PAP for adult cockerels,
based on ash and fat content of PAP (CVB, 2005):
AMEn (MJ) = (14,200 − 19.15 × ash content
+ 2.51 × fat content)/1,000. [3]
Based on this equation, calculated AMEn values
showed a good relationship with the determined AMEn
Downloaded from http://ps.oxfordjournals.org/ at University of Georgia on June 6, 2015
values in the current study (P < 0.001; R2 = 81.5). The
R2 could probably have been increased further if adult
cockerels had been used in the current study.
In the current study, a linear negative relationship
was found between ash content and protein content (P
< 0.001, R2 = 88.2). The effect of ash content on the
nutritional value of protein is not clear from the litera-
ture. An increase in ash content is expected to affect
the nutritional value of protein negatively because of
the changes in amino acid concentrations and amino
acid digestibility. Indeed, the protein efficiency ratio
(chick BW gain per unit of CP intake) decreased from
1.7 to 1.0 as ash content increased from 24 to 35% in 2
samples of meat and bone meal (Johnson and Parsons,
1997; Johnson et al., 1998). Mean amino acid digest-
ibility values of the 24 and 35% ash meat and bone
meal samples, however, were not significantly different
(P > 0.05; 70.8 and 76.3%, respectively), showing that
increased bone ash had no negative effect on amino
acid digestibility in the 2 meat and bone meal samples
evaluated in previous studies. In addition, in the cur-
rent study, digestibility coefficients of CP of the PAP
were not affected by ash content (P = 0.739). Shirley
and Parsons (2001) concluded that the negative rela-
tionship between protein quality and ash content of
meat and bone meal is almost entirely due to negative
effects on amino acid balance or profile of the meat and
bone meal, and not because of reduced amino acid di-
gestibility. In a study with broilers, however, digestibil-
ity of amino acids, with the exception of aspartic acid,
threonine, serine, tyrosine, histidine, and cysteine, was
negatively correlated with ash content, with samples
with high ash levels having lower digestibility (Ravin-
dran et al., 2002).
Differences in pressure during the processing of PAP,
however, might result in differences in amino acid di-
gestibility. Shirley and Parsons (2000) showed that
true digestibility values of most amino acids, particu-
larly of cysteine and lysine, were significantly decreased
with increasing pressures. It is hypothesized that the
relatively low cysteine digestibility of Sonac-60 (55%),
2614 van Krimpen et al.
Table 4. Digestibility coefficients (%, mean ± SD) of DM, ash, organic matter, CP, fat, gross energy, AMEn (kcal/kg), and amino
acids of the processed animal protein sources used in experiment 1 and 21
Meat and bone meal Meat meal

Item Daka-40 Sonac-50 Daka-58 Sonac-60

nutrient2
Digestibility coefficient of
  DM (%) 38.42 ± 7.76 55.74 ± 4.31 52.63 ± 3.67 64.28 ± 6.26
  Organic matter (%) 47.10 ± 8.69 64.63 ± 3.41 53.65 ± 4.99 61.33 ± 6.76
  CP (%) 81.43 ± 3.42 90.06 ± 1.93 85.26 ± 1.05 83.37 ± 1.54
  Fat (%) 79.65 ± 8.02 81.20 ± 2.32 94.49 ± 3.80 97.20 ± 6.18
  Gross energy (%) 55.95 ± 7.40 66.61 ± 2.81 62.01 ± 2.53 65.44 ± 5.58
  AMEn (kcal/kg) 1,819 2,566 2,801 3,107
Digestibility coefficient of amino acid2 (%)
  Lysine 78.1 ± 3.80 91.5 ± 1.18 81.0 ± 2.52 86.2 ± 3.69
  Methionine 75.3 ± 4.23 86.0 ± 1.71 79.9 ± 1.25 82.7 ± 1.81
  Cysteine 41.4 ± 12.27 88.4 ± 2.78 64.5 ± 7.29 54.8 ± 6.07
  Methionine + cysteine 67.3 ± 5.80 86.7 ± 1.99 76.1 ± 2.62 72.5 ± 3.35
  Threonine 68.0 ± 5.29 84.8 ± 2.17 77.8 ± 5.96 79.2 ± 3.92
  Tryptophan 67.0 ± 7.43 90.7 ± 2.12 71.5 ± 3.18 72.5 ± 7.58
  Isoleucine 67.9 ± 3.26 87.7 ± 3.74 77.1 ± 1.78 76.8 ± 4.24
  Arginine 87.2 ± 3.73 94.2 ± 1.41 91.5 ± 0.69 87.5 ± 2.35
  Phenylalanine 76.0 ± 6.30 90.8 ± 4.02 80.1 ± 2.92 81.9 ± 2.31

Downloaded from http://ps.oxfordjournals.org/ at University of Georgia on June 6, 2015

  Histidine 77.3 ± 3.44 92.6 ± 0.87 82.4 ± 3.81 83.2 ± 4.87
  Leucine 74.4 ± 6.20 89.7 ± 2.99 79.3 ± 1.16 78.8 ± 2.62
  Tyrosine 72.9 ± 8.36 92.2 ± 2.96 77.7 ± 6.08 79.4 ± 5.86
  Valine 71.7 ± 4.71 88.6 ± 2.87 76.8 ± 1.36 76.7 ± 1.89
  Alanine 68.5 ± 5.31 83.0 ± 5.10 71.0 ± 2.08 73.1 ± 2.06
  Aspartic acid 63.7 ± 3.73 86.3 ± 1.35 70.1 ± 5.96 75.5 ± 5.92
  Glutamic acid 78.9 ± 3.88 90.7 ± 0.74 81.3 ± 3.70 82.6 ± 2.57
  Glycine 62.0 ± 3.65 68.1 ± 4.33 58.7 ± 5.12 61.3 ± 2.11
  Proline 81.6 ± 5.33 91.1 ± 1.62 85.8 ± 4.43 84.9 ± 3.88
  Serine 77.9 ± 4.15 90.7 ± 0.54 83.3 ± 3.90 80.7 ± 3.56
  Sum amino acids 73.3 ± 3.94 86.4 ± 1.57 76.5 ± 2.29 77.7 ± 2.82
1Sonac-50 and Sonac-60 were provided by Sonac BV (Son, the Netherlands), and Daka-40 and Daka-58 were provided by Daka Proteins (Ringsted,
Denmark).
2Each value is the mean of 4 observations.

Daka-40 (41%), and Daka-58 (65%), compared with
Sonac-50 (88%), was partly caused by high pressure
during processing.
Effect of PAP Sources on Performance
Feed intake level of the 2 meat meal diets (Sonac-60
and Daka-58) was higher compared with the intake
level of the 2 meat and bone meal diets (Sonac-50 and
Daka-40). Nutritional values of the PAP were deter-
mined in the digestibility study, and these values were
used in calculating the diets for the performance study.
Therefore, it was assumed that the nutritional values of
the different diets were similar. Hence, these differences
in feed intake were not expected because it is generally
known that laying hens adjust their feed intake to their
nutritional needs (van Krimpen et al., 2007, 2008, 2009;
Van der Meulen et al., 2008). It is unknown which spe-
cific property in the meat and bone meals was respon-
sible for the reduced feed intake levels.
Laying hen performance differed between treatments;
performance was most favorable for the Sonac-50 treat-
ment and most adverse for the Daka-40 treatment. Al-
though contents of the digestible essential amino ac-
ids were equated for all diets, amino acid intake levels
differed between treatments due to differences in feed
intake level. Because of the different feed intake levels
and different dietary contents, intake levels of the non-
essential amino acids also differed between treatments.
Egg mass production was probably partly affected by
different intake levels of (non)essential amino acids. Sig-
nificant positive linear regressions were found between
intake of digestible aspartic acid (R2 = 0.19), cysteine
(R2 = 0.18), glutamic acid (R2 = 0.16), histidine (R2
= 0.21), isoleucine (R2 = 0.16), leucine (R2 = 0.12),
phenylalanine (R2 = 0.13), serine (R2 = 0.15), tyrosine
(R2 = 0.14), valine (R2 = 0.14), and egg mass produc-
tion (g/hen per d). Compared with the control diet, egg
weights of the PAP-rich diets were similar (Sonac-60)
or (slightly) reduced (Sonac-50, Daka-40, Daka-58).
Several other studies showed reduced egg weights in
PAP-fed hens (Damron et al., 2001; Bozkurt et al.,
2004). This effect might be associated with the amino
acid concentration of the PAP used (Parsons et al.,
1997; Wang and Parsons, 1998b; Shirley and Parsons,
2001). In the current study, however, differences in egg
weights could only partially be explained by variations
in the intake of digestible amino acids. Significant posi-
tive linear regressions were found for cysteine (R2 =
0.07), glutamic acid (R2 = 0.05), histidine (R2 = 0.04),
and serine (R2 = 0.05), but levels of declared variance
were very low.
Nutritional factors, such as dietary protein and min-
eral contents, could affect litter conditions (Veldkamp
et al., 2007; Enting et al., 2009). In our study, we found
significant linear regressions between litter wetness and
 VEGETABLE VERSUS ANIMAL PROTEIN IN LAYER DIET 2615
intake of calcium (R2
= 0.46), chloride (R2
= 0.36), po-
tassium (R2 = 0.56), sodium (R2 = 0.46). Intake of CP

Treatment × week
showed a weak relationship with litter wetness (R2 =

0.321
0.939
<0.001
0.998
0.998
0.758
0.427
0.10). Compared with the hens fed Sonac-60, only the
hens fed meat and bone meal (Daka-40 and Sonac-50)
showed significantly lower intake levels of calcium
(4,730 vs. 4,524 and 4,520 mg/hen per d), potassium
(754 vs. 703 and 705 mg/hen per d), and sodium (174
P-value

<0.001 vs. 167 mg/hen per d).

<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
Week

From this study, it can be concluded that nutritional

values of the tested PAP sources varied wideely. De-
spite the fact that the diets were balanced for these

and Sonac-60 were provided by Sonac BV (Son, the Netherlands), and Daka-40 and Daka-58 were provided by Daka Proteins (Ringsted, Denmark).
differences, feed intake levels of hens fed the meat and
Treatment

0.005
0.163
0.014
0.001
0.049
0.156
<0.001 bone meal diets were reduced compared with those of
hens fed the meat meal diets, whereas the feed intake
level of hens fed the control diet was intermediate. Dif-
ferences in feed intake and subsequent mineral intake
seemed to be related to differences in litter wetness.
1.509
1.214
0.595
0.754
0.058

Downloaded from http://ps.oxfordjournals.org/ at University of Georgia on June 6, 2015

20.78
3.07
SE

Laying hen performance differed between treatments,

being most favorable for the Sonac-50 treatment and
most adverse for the Daka-40 treatment. Differences in
laying hen performance seemed to be partially related
2.30a
Sonac-60

48.4ab
124.5a

32.5a
77.6
59.9

to differences in feed intake and corresponding amino

1,788

acid intake.
Meat meal

ACKNOWLEDGMENTS
2.26ab
Daka-58

122.3ab

47.6bc

32.4a
78.2
58.4

1,781

The authors gratefully acknowledge funding from the

Table 5. Performance traits per treatment in ISA Brown laying hens from 21 to 40 wk of age1

Product Board for Livestock and Eggs (Zoetermeer,

the Netherlands), Daka Proteins (Ringsted, Denmark),
Vion Food (Son, the Netherlands), and EFRA (Brussels,
Belgium). We also thank Remco de Veer (Wageningen
2.13c
Sonac-50

24.8b
49.2a
118.9c
79.7
59.4

University, Wageningen, the Netherlands), Gisabeth

Meat and bone meal

1,795

within a row with different superscripts are significantly different (P < 0.05).

Binnendijk (Wageningen UR, Lelystad, the Nether-

lands), and the poultry keepers of the accommodation
De Haar (Wageningen University) for their enormous
2.22abc

help in performing all observations.

Daka-40

21.1b
119.1c

46.4c
76.5
58.7

1,767

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