Anais Completo

IV INTERNATIONAL SYMPOSIUM ON
NUTRITIONAL REQUIREMENTS
OF POULTRY AND SWINE
ANAIS
Editors:
Luiz Fernando Teixeira Albino

Horacio Santiago Rostagno
March 29 and 30, 2017

Viçosa – MG – Brazil
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UNIVERSIDADE FEDERAL DE VIÇOSA

DEPARTAMENTO DE ZOOTECNIA
IV INTERNATIONAL SYMPOSIUM ON
March 29 and 30, 2017

iv
Copies of this book can be purchased at:

Departamento de Zootecnia
Universidade Federal de Viçosa
Viçosa – MG – 36570-000
(0xx) 31-3899-3310
Editors:
Horacio Santiago Rostagno
Editoring:
Edson Agostinho Pereira (0xx) 31-3899-2677
Catalog file prepared by Catalogation Section and Classification

of the Central Library of UFV
I61 International Symposium on Nutritional Requirement of Poultry

2917 and Swine (4. : 2017 : Viçosa, MG)
Anais [do] 4. International Symposium on Nutritional
Requirement of Poultry and Swine, 29 and 30 march 2017.
Viçosa, MG ; Editors Luiz Fernando Teixeira Albino, Horacio
Santiago Rostagno . – Viçosa, MG : UFV, DZO, 2017.
xii, 374p. ;
Inclui referências.
ISBN: 978-85-8179-123-4
1. Nutrição animal - Congressos. 2. Aves – Alimentação e

Rações. 3. Suínos – Alimentação e rações. I. Albino, Luiz
Fernando. II. Rostagno, Luiz Fernando Teixeira. III.
Universidade Federal de Viçosa. Departamento de Zootecnia.
IV. Título.
CDD 636.085
v
III INTERNATIONAL SYMPOSIUM ON

PROCEEDINGS
Organizing Committee
Horacio Santiago Rostagno (Chairman)
Melissa Izabel Hannas
Sandra Carolina Salguero Cruz
Valdir Ribeiro Junior
Edson Agostinho Pereira
Bruno Reis de Carvalho
Rosana Cardoso Maia
João Paulo de Oliveira
March 29 and 30, 2017

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vii
SPECIAL THANKS TO THE COMPANIES FOR

THE FINANTIAL SUPPORT
ABVISTA
ADISSEO
AGROCERES MULTIMIX
AJINOMOTO
ALLTECH
BIOMIN
BTECH
CARGILL
COGRAN
DSM
EVONIK
IMPEXTRACO
NUTRIQUEST TECHNOFEED
PHILEO
POLINUTRI
VACCINAR
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Preface
In Brazil research with domestic animals mainly poultry and
swine has become relevant in the last three decades and scientific
studies in the area had reduced the dependence on foreign
literature.
In 1996 was held in Viçosa the first International
Symposium that resulted in the publication of a significant amount
of information about the nutritional requirements of poultry and
swine, which was a useful tool for teachers and the feed
compound industry. The II and III International Symposium held in
2005 and 2011 with the join presentation of the 2 nd and 3rd edition
of the Brazilian Tables for Poultry and Swine were of special
importance for planning new lines of research and improving the
formulation of diets for poultry and swine.
Today with the realization of the IV International
Symposium, the most recent nutrition results of investigations,
from Brazil and abroad, are presented.
Considering that the two main objectives of the symposium
are first, to discuss the current systems of nutritional requirements
techniques and second, to promote a strong relationship among
nutritionists of the industries and researchers of the area. It is
clearly understood that the acquired knowledge during this
symposium and the information contained in this publication will
make possible an improvement in the production index of poultry
and swine.
Luiz Fernando Teixeira Albino and Horacio Santiago Rostagno

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ÍNDICE
FEEDSTUFF COMPOSITION FOR POULTRY AND SWINE….. 1

Markus Karl Wiltafsky, Athanasios Tsakonas, Jaroslav Heger, Jiri Zelenka
SIMULATION MODELS: USE OF ENERGY BY POULTRY ….. 33
Nilva Kazue Sakomura, Nayara Tavares Ferreira, Rob Mervyn Gous
AMINO ACIDS IN BROILER NUTRITION: ALIGNING PRACTICE
WITH SCIENCE………………………………………………………… 69
Gabriel B. S. Pessoa, Eduardo T. Nogueira, Marianne Kutschenko, Luciana
F. de Lima
NUTRITIONAL RECOMMENDATIONS FOR LAYING HENS

AND JAPANESE QUAILS…………………………………………… 99
Fernando Guilherme Perazzo Costa, Matheus Ramalho de Lima, Danilo
Cavalcante, Guilherme Souza Lima, Danilo Vargas Gonçalves Vieira
INORGANIC AND ORGANIC TRACE MINERALS NUTRITION
OF BROILERS………………………………………………………… 123
Ramon D. Malheiros and Peter R. Ferket
RECOMMENDED VITAMIN LEVELS FOR HIGH
PERFORMANCE BROILERS AND LAYERS……………………… 131
Gilberto Litta
NUTRITIONAL REQUIREMENTS OF AMINO ACIDS FOR
GROWING PIGS……………………………………………………… 181
John K. Htoo
VALIDATION OF THE NUTRITIONAL REQUIREMENT FOR
HIGH PERFORMANCE SWINE……………………………………. 209
Uislei Orlando, Márcio Gonçalves, Wayne Cast, Matthew Culbertson, and
Keysuke Muramatsu
NUTRITIONAL REQUIREMENTS OF AMINO ACIDS FOR
PIGLET………………………………………………………………… 243
Etienne Corrent and Aude Simongiovanni
xii
AMINO ACIDS ESSENTIALITY FOR SOWS: NEW APPROACHES 273

Márvio Lobão Teixeira de Abreu, Alysson Saraiva
AMINO ACIDS: A TOOL TO IMPROVE MEAT QUALITY……… 291
Pierre-André Geraert
BASIC CONCEPTS OF STATISTICS THAT A NUTRITIONIST
MUST 315
KNOW………………………………………………………………….
Fabyano Fonseca e Silva
STRATEGIES TO INCREASE PIGLETS PERFORMANCE
AFTER WEANING …………………………………………………… 341
José Francisco Pérez, Laia Blavi and David Solá-Oriol
FEEDSTUFF COMPOSITION FOR POULTRY AND
SWINE
1 1 2 2
Markus Karl Wiltafsky , Athanasios Tsakonas , Jaroslav Heger , Jiri Zelenka
1
Evonik Nutrition & Care GmbH, Rodenbacher Chaussee 4, 63457 Hanau, Germany
2
Department of Animal Nutrition and Forage Production, Mendel University in Brno,
613 00 Brno, Czech Republic;
ABSTRACT
A sufficient heat-treatment of soybeans is necessary in

order to inactivate the antinutritonal factors which naturally occur
in soybeans. At the same time, over-processing needs to be
avoided. A growth trial was conducted to investigate the effect of
different heat treatments imposed on full-fat soybeans on the
performance and organ weights in broilers. This experiment was
conducted with 5040 day-old male Ross 308 chicks and 10 dietary
treatments with 6 replicates per treatment. Treatments (T) 1 and
10 had 140 chicks per replicate and Treatments 2 to 7 had 70
chicks per replicate. Diets were based on wheat (23.8 – 36.5%)
and corn (20.0 – 35.0%) and contained full-fat soybeans or
soybean meal as major protein source. Dietary energy levels were
in line with Ross 308 recommendations (2007). Calculated
standardized ileal digestible (SID) lysine levels were adequate in
T8 and T9, and set at 95% of the recommended level in the other
treatments. The diets of T1 contained raw soybeans (negative
control; Starter 40%; Grower 39%; Finisher 34%), and the diets
from T2 to T7 contained the same amount of differently processed
full-fat soybeans whereas T8 contained higher amounts of full-fat
soybeans in the Grower and Finisher feed (Grower 43%; Finisher
37%). T9 and T10 were positive control groups containing
soybean meal (Starter 37.2 and 34.6%; Grower 31.1 and 28.5%;
Finisher 26.9 and 24.6%, respectively), with lysine set at 100%
and 95% of the recommendation, respectively. Trypsin inhibitor
activity of full-fat soybeans decreased as processing time and/or
temperature increased. The best results in terms of feed intake,
2 - IV International Symposium on Nutritional Requirements of Poultry and Swine
growth rate, feed conversion ratio, carcass and breast meat yield
were obtained if the full-fat soybeans were short-term (1 min,
100°C) and long-term (5 min, 100°C) conditioned and expanded
for 15 sec at 125°C (T6), resulting in a TIA level of 5.9 mg/g.
Concurrently, performance of chickens improved from T1 to 6 (P <
0.05). At day 35, weight gain and feed conversion ratio of T6 were
improved by 59% and 22% compared to T1, respectively.
Numerically better performance was found in 100 % amino acid
diets as compared with 95 % diets (P > 0.05), for full-fat soybeans
(T8 vs T7) and for SBM containing diets (T9 vs T10). The
performance achieved with the most intensively processed full-fat
soybeans was at the same level as that achieved with SBM,
although the TIA level of 5.3 mg/g for this full-fat soybeans was
higher than the commonly recommended maximum level of 4.0
mg/g (95% AA level: T7 vs T10; 100% AA level: T8 vs T9).
In this trial it was shown that today’s broilers are able to
achieve good performances if they are fed with adequately
processed full-fat soybeans as the main protein source. Broilers
respond to an under-supply with amino acids even if the amino
acid level is just 5% below the recommended levels.
In a second experiment, the standardized ileal digestibility
of amino acids was estimated in the full-fat soybeans qualities
used in the growth trial and in 9 additional full-fat soybeans
qualities (Z1 - Z9) which were sub-batches of the full-fat soybeans
used in T8 but further processed in an autoclave to provoke over-
processing. Diets were calculated to be adequate in energy,
minerals and vitamins and contained full-fat soybeans as the only
source of amino acids. Each diet was offered ad libitum to six
cages (6 birds/cage) of male Ross 308 broilers from 25 to 28 days
of age. Pooled ileal digesta samples were collected on day 28
from all birds within one cage.
Low amino acid digestibility coefficients were recorded in
raw soybeans (T1) and digestibility improved with increasing
processing intensity. However, as processing prolonged the
digestibility decreased again. For example the standardized ileal
IV International Symposium on Nutritional Requirements of Poultry and Swine - 3
digestibility of Lys increased from 50.8% in T1 to 84.7% in Z2 and

was reduced down to 39.8% in Z9.
In conclusion, there is a statistically significant impact
(99.9% confidence level) of processing on the amino acid
digestibility and under- as well as over-processing will reduce it.
INTRODUCTION
Today’s livestock production is in the focus of quite different

discussions. It shall deliver high quality food to cover the ever
growing demand of the human mankind. At the same time, this
target shall be achieved with shrinking resources which is only
possible if the overall efficiency of livestock production is
increased. The improvements via breeding are quite impressive.
For example, body weight of broilers at day 42 improved from
1957 to 2001 from 578g to 2672g and feed conversion ratio
improved from 2.14 to 1.62 (Havenstein et al., 2003). It is obvious
that these changes in the genetic potential require continuous
updates of the nutritional recommendations. For example, in 1998
the standardized ileal digestible (SID) Lys recommendation for 10-
20 kg pigs was 1.01% (NRC, 1998) whereas it increased to 1.23%
in 2012 (NRC, 2012) and to 1.32% in 2016 (AMINODat 5.0). A
good knowledge of the animal’s amino acid needs is essential to
avoid safety margins and amino acid surpluses. Thus, it is the
basis for optimization of diet composition which increases feed
efficiency and reduces the environmental impact of animal
husbandry. The use of essential amino acids in feed formulation
decreases demand for protein sources. Supplementation with
essential amino acids enables feeding of low-protein diets and
usage of protein sources of minor quality because imbalances in
the amino acid pattern can be corrected. Prevention of surface
and ground water contamination with nitrogen is achievable by
reductions of crude protein (CP) and is desirable for both
economic and environmental reasons. On average, a 10%
reduction in total nitrogen excretion was reported for every 1%
reduction in CP content of the diet (Kerr and Easter, 1995; Le
Bellego and Noblet, 2002; Shriver et al, 2003; Lordelo et al.,

2008).
The target of increased efficiency in livestock production
can only be achieved if the diet formulation is based on the correct
matrix values. Updating matrix values of feed ingredients is an
ongoing task which requires continuous analyses. The
developments in near infrared reflectance (NIR) technology offer
feed millers a valuable tool to solve this task. In the meantime NIR
calibrations are available not only for proximate parameters like
crude protein and fat content but also for amino acids. It was well
described for example by Ravindran et al. (2014) how variable the
quality of soybean meal (SBM) is. This is not only true for the total
content of nutrients but also for their digestibility. Especially the
impact of processing plays a major role in causing qualitative
differences between raw materials.
In the last years, the feed industry showed a growing
interest in the use of full-fat soybeans (FFSB) and, as a result, its
large-scale processing became a common practice. Due to their
high oil content, FFSB are particularly suitable for manufacturing
high-energy poultry diets, as post-pellet application of fat may be
reduced or eliminated (Waldroup, 1982). However, the nutritional
potential of FFSB is limited by the presence of antinutritional
factors, mainly trypsin inhibitors, which interfere with digestion,
absorption and metabolism of nutrients (Liener and Kakade,
1980). Trypsin inhibitors form complexes with pancreatic
proteases, thus reducing their acitivity in the upper small intestine.
In an attempt to offset the lack of enzymes, hypersecretion by the
exocrine pancreas is evoked followed by pancreatic hypertrophy
and hyperplasia (Grant et al., 1995). As a result of the combined
effect of endogenous loss of essential amino acids (particularly
sulphur amino acids) and decreased intestinal proteolysis, growth
performance of chickens is reduced.
Thus, sufficient inactivation of trypsin inhibitors is crucial for
optimal broiler performance. However, over-processing must be
avoided to prevent the reduction of the nutritional value of the
product. Excessive heat treatment may result in decreased amino

acid availability due to the early Maillard reaction. Lysine is
particularly susceptible because of its exposed ε-amino group
which readily reacts with reducing sugars. During the advanced
phase of Maillard reaction, cross-linking may occur between most
amino acids and polypeptide chains, thus reducing the efficiency
of intestinal proteases. Part of amino acids may be totally
destroyed as a result of advanced Maillard reaction. Thus, an
optimal balance between under- and over-processing must be
found to ensure an efficient utilization of soybean protein. In the
first experiment, an attempt was made to find such optimum and to
investigate which amount of trypsin inhibitors was tolerated by
growing broilers without affecting their performance. In a second
experiment, the impact of processing on the amino acid
digestibility in broilers was investigated. The aim of the present
experiments was to investigate the effect of different heat
treatments imposed on FFSB on the performance and organ
weights in broilers and the amino acid digestibility.
Table 1 - Processing conditions used to produce sub-batches of

full-fat soybeans differing in their quality
Conditioning Autoclaving
Expanding at 110 °C
Soybean Short-term Long-term (15 s)
Treatment (60 s)
source
o o o
C min C C min
T1 FFSB - - - - -
T2 FFSB 80 - - 115 -
T3 FFSB 100 5 100 115 -
T4 FFSB 100 15 100 115 -
T5 FFSB 80 - - 125 -
T6 FFSB 100 5 100 125 -
T7 FFSB 100 15 100 125 -
T8 FFSB 100 15 100 125 -
T9 SBM - - - - -
T10 SBM - - - - -
Z1 FFSB 100 15 100 125 15
Z2 FFSB 100 15 100 125 30
Z3 FFSB 100 15 100 125 45
Z4 FFSB 100 15 100 125 60
Z5 FFSB 100 15 100 125 120
Z6 FFSB 100 15 100 125 180
Z7 FFSB 100 15 100 125 240
Z8 FFSB 100 15 100 125 300
Z9 FFSB 100 15 100 125 360
FFSB - full-fat soybeans.

Processing of soybeans – T1 to T8
One batch of broken, raw soybeans was purchased from

Rieder Asamhof GmbH & Co. KG (Kissing, Germany). The
following equipment had been used for the processing of the
soybeans: short-term conditioner (Amandus Kahl GmbH & Co.
KG, Reinbeck Germany, Typ: DLM I), long-term conditioner
(Amandus Kahl GmbH & Co. KG, Reinbeck, Germany, Typ: LK
1605-2) and expander (Amandus Kahl GmbH & Co. KG, Reinbeck
Germany, Typ: OEE 15.2). Various combinations of short-term
conditioning at 80 or 100oC, long term conditioning at 100oC for 5
or 15 min and expanding at 115 or 125oC were applied to
manufacture six FFSB products used in experimental diets (Table 1).
The expander had a throughput of 1.5 to 1.8 ton per hour. After
short-term conditioning at 80°C, a specific energy input of 30.0
and 39.2 kWh/ton was needed to reach expansion temperatures of
115 and 125°C, respectively. After short-term conditioning at
100°C and long-term conditioning at 100°C for 5 and 15 min, a
specific energy input of 13.5 and 20.0 kWh/ton was needed to
reach expansion temperatures of 115 and 125°C, respectively.
All FFSB products were dried in a belt dryer with 85°C air
and left the dryer after 5 min with a temperature of 43°C. Then
they entered the belt cooler and left it after 5 min with a
temperature of 30°C.
Processing of soybeans – Z1 to Z9
Sub-batches of T8 were further processed at 110°C over

different times lasting from 15 to 360 min using an autoclave (Typ
HST 6x9x12, Zirbus Technology GmbH, Bad Grund, Germany).
The autoclave worked under an automatic mode to minimize the
human impact. The settings of the warming-up and cooling-down
phase were the same for all treatments. During the cooling-down
phase multiple vacuum phases were applied to reduce the final
moisture content back to about 12% and to guarantee storage
stability of the final product. The detailed processing conditions are

shown in Table 1.
Animals and Procedures – Growth trial
The experiment was conducted at the International Poultry

Testing Station Ústrašice, Czech Republic. The animal procedures
were reviewed and approved by the Animal Care Committee of the
Mendel University in Brno. A total of 5040 one-day-old male Ross
308 broiler chicks were allocated randomly to 10 dietary
treatments using a randomized complete block design with six
replicates per treatment. There were 70 chicks per replicate pen
except for treatments 1 and 10, in which 140 chicks per pen were
used. The chickens were kept in a windowless house with full
climatic control, on deep litter from wood shavings. Each pen was
equipped with manually filled tube feeders and nipple drinkers.
Heating and lighting programs were in accordance with Ross
Broiler Management Manual (2009). On day 1, the birds in each
pen were weighed together while on days 10, 24 and 35, all birds
were weighed individually. At the same time, feed consumption
per pen was recorded. On d 35, five birds of each pen having body
weights closest to the pen mean were selected, tagged and after
twelve hours of fasting they were weighed, sacrificed and carcass,
breast meat (boneless and without skin) and pancreas weights
were recorded.
Diets – Growth trial
Starter (d 1‒10), grower (d 11-24) and finisher (d 25 ‒ 35)

diets were formulated to contain FFSB or SBM as main protein
sources. Except for amino acids, the isocaloric diets were
formulated to be in line with the Ross nutrition supplements
(2009). To enable sensitive detection of changes in soybean
protein quality, the diets for T1 to T7 and T10 were calculated to
contain standardized ileal digestible amino acids at levels
corresponding to 95 % of the recommended requirement. The diet

containing raw FFSB served as a negative control (T1). Various
combinations of processing conditions were applied to
manufacture six further FFSB products used in diets of the same
composition as the control diet (T2 to T7). T8 was the same as T7
except that the amino acid level was increased to 100 % of the
requirement. Diets for T9 and T10 were positive controls using
commercial SBM as the main protein source and containing amino
acids at levels of 95 and 100 %, respectively. Processing
conditions are summarized in Table 1. Ingredient composition and
nutrient contents of diets are given in Table 2 and Table 3. The
diets in crumbled form (starter phase) or pellets (grower and
finisher phase) were supplied ad libitum.
Table 3 - Nutrient content of the diets used in the growth trial (as-fed basis)
Diet Starter (0-10d) Grower (11-24d) Finisher (25-35d)
Treatment T1-7 T8 T9 T10 T1-7 T8 T9 T10 T1-7 T8 T9 T10
Amino acid level % 95 100 100 95 95 100 100 95 95 100 100 95
Calculated composition
AMEn (kcal/kg) 3025 3025 3025 3025 3150 3150 3150 3150 3200 3200 3200 3200
Crude protein (g/kg) 225.0 234.4 233.8 224.8 199.3 208.4 207.4 198.4 182.9 190.8 189.9 182.2
Crude fat (g/kg) 87.0 87.2 70.4 67.3 91.0 95.4 79.9 76.8 91.3 95.1 81.7 79.0
SID Lysine (g/kg) 12.1 12.7 12.7 12.1 10.4 11.0 11.0 10.4 9.2 9.7 9.7 9.2
SID Met+Cys (g/kg) 8.7 9.1 9.1 8.7 7.7 8.1 8.1 7.7 7.0 7.4 7.4 7.0
SID Threonine (g/kg) 7.6 8.0 8.0 7.6 6.7 7.0 7.0 6.7 6.0 6.3 6.3 6.0
SID Valine (g/kg) 9.6 10.0 10.0 9.6 8.3 8.8 8.8 8.3 7.4 7.8 7.8 7.4
SID Isoleucine (g/kg) 8.2 8.6 8.6 8.2 7.2 7.6 7.6 7.2 6.5 6.9 6.9 6.5
Analysed composition
Crude protein (g/kg) 230.2 240.3 238.9 230.7 199.3 206.1 212.6 203.5 182.9 194.1 197.8 183.8
Lysine (g/kg) 13.6 14.3 13.6 13.2 11.8 12.5 12.2 11.6 10.4 11.2 11.2 9.9
Methionine (g/kg) 5.8 6.2 5.9 5.6 5.1 5.2 5.1 4.7 4.6 4.9 4.8 4.2
Cysteine (g/kg) 3.8 4.0 3.7 3.5 3.7 3.8 3.6 3.7 3.3 3.4 3.3 3.2
Threonine (g/kg) 8.9 9.6 9.0 8.9 7.7 8.1 8.1 7.8 7.2 7.6 7.5 6.9
Arginine (g/kg) 15.3 16.2 15.9 15.4 12.6 13.3 13.7 13.1 11.8 12.6 13.0 11.6
Valine (g/kg) 10.8 11.5 11.4 10.9 9.4 9.8 10.0 9.6 8.5 9.2 9.4 8.4
Isoleucine (g/kg) 9.3 9.8 9.8 9.5 8.2 8.6 8.8 8.5 7.6 8.1 8.3 7.4
SID - standardized ileal digestible.

IV International Symposium on Nutritional Requirements of Poultry and Swine -
11
Animals and Procedures – Digestibility trial
The digestibility trial was conducted in cooperation with

Prof. Ravindran at Massey University, New Zealand. A total of 16
assay diets (Table 4) based on FFSB (16 different qualities; Table
1) as the only source of protein were formulated to supply
approximately 18% crude protein in the diet. Diets were calculated
to be adequate in energy, minerals and vitamins. The diets
contained titanium dioxide as an indigestible marker.
Table 4 - Dietary composition of the assay diets

Ingredient Inclusion rate, %
full-fat soybeans 50.00
Dextrose 44.10
Soybean oil 2.00
Titanium dioxide 0.30
Sodium bicarbonate 0.20
Dicalcium phosphate 1.90
Limestone 1.00
Trace mineral premix 0.25
Vitamin premix 0.05
Salt 0.20
Each diet was offered ad libitum to six cages (6 birds/cage)

of male Ross 308 broilers from 25 to 28 days of age. On day 28,
all birds were euthanised by an intracardial injection of diluted
sodium pentobarbitone solution, and the contents of the lower half
of the ileum was collected by gently flushing with distilled water
into plastic containers (Ravindran et al., 2005). The ileum was
defined as that portion of the small intestine extending from
Meckel's diverticulum to a point 40 mm proximal to the ileocaecal
junction. Samples from all birds within one cage were pooled,
frozen immediately after collection and subsequently freeze-dried.
Ingredient, diet and ileal digesta samples were ground to
pass through a 0.5 mm sieve and stored in airtight containers at -
20 °C for chemical analyses (dry matter, titanium, nitrogen, and

amino acids). Amino acid and nitrogen analyses were performed
by Evonik Nutrition & Care GmbH and determination of titanium
was performed by Massey University, New Zealand.
Calculations
Apparent ileal nitrogen and amino acid digestibility

coefficients were calculated from the dietary ratio of amino acid to
titanium relative to the corresponding ratio in the ileal digesta.
Apparent ileal digestibility coefficient (AID, %) = ((AA / Ti) d - (AA

/ Ti)i) / (AA / Ti)d x 100
Where, (AA / Ti)d = ratio of amino acid to titanium in diet and (AA /
Ti)i = ratio of amino acid to titanium in ileal digesta.
Apparent digestibility data were converted to standardized

values, using endogenous AA values published by Lemme et al.
(2004). The following formula shows the calculation of
standardised ileal digestibility (SID) of an amino acid in the test
ingredient.
SID (%) = (AID (%) + [Basal eAA losses (g/kg DMI)] x 100) /
Ingredient AA (g/kg DM)
Where, AID = apparent ileal digestibility coefficient of the amino acid

Basal eAA losses= Basal endogenous amino acid losses
Ingredient AA = Concentration of the amino acid in the ingredient
Chemical Analyses
The raw and processed FFSB as well as commercial SBM

were analyzed for urease activity (UA) according to ISO
5506:1988, protein solubility in a 0.2% potassium-hydroxide
solution (PS) as described by Araba and Dale (1990a), protein
dispersibility index (PDI) using the method of the American
Association of Cereal Chemists (AOCS 1996), and trypsin inhibitor

activity (TIA) according to ISO 14902:2001. Reactive lysine was
determined in raw and FFSB samples using homoarginine
reaction as described by Fontaine et al. (2007). The diets and
digesta samples were analyzed for nitrogen using Dumas
procedure and for amino acids by ion-exchange chromatography
(Llames and Fontaine, 1994). The concentration of titanium was
determined using an inductively coupled plasma optical emission
spectrometer following a sulphuric acid and nitric acid wet
digestion (Boguhn et al., 2009).
Statistical Analysis
Experimental data were analyzed as a completely

randomized block design using ANOVA procedure of
Statgraphics Plus package (version 3.1., Statistical Graphis
Corp., Rockville, MD). When a significant value for treatment
effect (P < 0.05) was observed, the differences between means
were assessed by Tukey HSD test. The experimental unit was a
replicate pen. Linear, quadratic or rectilinear (Robbins et al.,
2006) models were fitted to experimental data to describe the
response of chickens to TIA values found in FFSB and SBM
samples. Only treatments with 95% amino acid levels were
included into this analysis. Pearson correlation coefficients
between in vitro indicators of soybean treatment as well as
between in vitro indicators and in vivo performance
characteristics were calculated.
The results of the digestibility trial were analyzed using
Multivariate ANOVA and 95% confidence intervals were derived
using t-test (R environment, version 3.3.2). For standardized
ileal digestibility coefficients of Lysine, a normality test for each
treatment was done using the Shapiro-Wilk test and the
Students t-test was done for each pair of treatments. In cases
where T6 and T7 are involved, the Kolmogorov-Smirnov test
was done, as it was shown that their data did not follow a
normal distribution. Hierarchical clustering of the treatments and

digestibility coefficients was done based on their similarity
(Euclidean distance).
RESULTS AND DISCUSSION
Growth Experiment
Except for PS all other in vitro indicators responded strongly

to changes in FFSB treatment (Table 5). Urease activity fell almost
to zero in T7 while PDI and TIA were reduced to 20 % of their
initial values. The lowest TIA of T1 to T7, was slightly above the
recommended threshold level of about 4.0 mg/g (Leeson et al..
1987; Loeffler et al.. 2012). The increase of the expanding
temperature from 115 to 125 oC (T2 vs. T5) resulted in a reduction
of PDI and TIA values by 57 and 70 %, respectively. The PS
gradually decreased from 94 to 89 %.
Table 5 - In-vitro indicators of soybean processing

Urease Protein Trypsin
Soya KOH protein
Treatment activity dispersibility inhibitor activity
source solubility (%)
(mg N/g) index (%) (mg/g)
T1 FFSB 4.15 72.2 94.2 27.3
T2 FFSB 3.88 50.7 99.0 24.0
T3 FFSB 1.53 34.6 93.3 14.4
T4 FFSB 1.19 27.1 91.9 8.7
T5 FFSB 0.42 22.0 90.0 7.2
T6 FFSB 0.36 24.6 89.5 5.9
T7 FFSB 0.05 14.7 88.7 5.3
T8 FFSB 0.05 14.7 88.7 5.3
T9 SBM 0.01 12.5 80.2 2.6
T10 SBM 0.01 12.5 80.2 2.6
Z1 FFSB 0.02 9.8 81.4 2.4
Z2 FFSB 0.01 7.5 71.8 1.5
Z3 FFSB 0.00 7.0 66.9 1.0
Z4 FFSB 0.00 6.8 60.3 0.8
Z5 FFSB 0.00 6.7 43.3 0.5
Z6 FFSB 0.00 6.9 30.2 0.4
Z7 FFSB 0.00 7.8 24.2 0.4
Z8 FFSB 0.00 7.7 19.4 0.3
Z9 FFSB 0.00 8.2 18.4 0.2
Although it has been suggested that the PS assay may be

useful in assessing protein quality in under-processed samples
(Araba and Dale, 1990b), it is commonly used as an indicator of
soybean over-processing (Parsons et al., 1991; Anderson-
Haferman et al., 1992). The critical level of PS associated with
maintaining optimal performance is a matter of debate. Lee et al.
(1991) reported that a 10 % drop in PS resulted in a significant
decrease in growth rate of turkeys while Araba and Dale (1990a)
and Parsons et al. (1991) identified the critical PS levels at 70 and
59 %, respectively. The PS values found in the present study thus
suggested that none of the FFSB treatment of T1 to T7 led to
soybean over-processing. As indicated by correlation coefficients

(Table 8), there was a close correlation between UA, PDI and TIA,
the r-values ranging between 0.97 and 0.99. A moderately strong
relationship was found between PS and other FFSB processing
indicators. Reactive lysine content in raw FFSB expressed as
percentage of total lysine was 92.4. The values found in other FFSB
samples ranged between 91.4 and 92.9, thus indicating no
deleterious effect of processing on lysine availability in T1 to T8.
Growth performance of chickens fed raw or under-processed
FFSB was markedly reduced in comparison with the SBM-fed birds
(Table 6). At 95 % amino acid level, both WG and FCR were
maximized in T6 and were not significantly different from those
obtained with SBM-based diets. Similar results were reported by
other authors comparing SBM and properly heat-treated FFSB
included into broiler diets at moderate levels (Herkelman et al.,
1991; Perilla et al., 1997; Mirghelenj et al., 2013). Increasing the
amino acid concentration from 95 % to 100 % of the requirement
resulted in an insignificant improvement of growth rate and FCR in
both types of diet. Feed intake followed a similar trend as growth
performance. In contrast. Perilla et al. (1997) reported a negative
relationship between extrusion temperature and feed intake in
chickens fed FFSB-based diets. Carcass and breast meat yields
responses to processing time and temperature were similar to those
of growth rate and FCR (Table 7). Relative pancreas weight was
reduced from 4.47 g/kg in raw FFSB (T1) to 1.85 g/kg in T3. In
other treatments, the values ranged between 1.73 and 2.26 g/kg
including birds fed SBM-based diets. The only exception was T5 in
which unexpectedly higher pancreas weights were found.
It is generally accepted that the reduction of chicken
performance and pancreatic hypertrophy is largely due to the
presence of trypsin inhibitors in raw soybeans (Applegarth et al.,
1964; Han and Parsons, 1991; Leeson and Attteh, 1996). In the
present study, a close correlation was found between in vivo
chicken response and in vitro indicators of FFSB treatment (Table
8). TIA and PDI were the best predictors of chicken growth
performance. Growth rate and FCR did not change appreciably at

TIA levels below 15 mg/kg which suggest that the TIA threshold
might be higher than the commonly accepted values of 4 to 5 mg/kg
(Leeson et al., 1987; Leeson and Atteh, 1996). PDI is considered to
be a sensitive indicator of minimum adequate heat processing of
SBM (Batal et al., 2000). The results of the present study suggest
that to attain sufficient heat treatment, the PDI value should not
exceed 30 to 35 %. A slightly higher threshold (45 %) was
suggested by Batal et al. (2000) while the American Soybean
Association recommended optimum PDI between 15 and 30 %
(Balloun, 1980).
In accordance with the results of other studies (Anderson-
Haferman et al., 1992; Ruiz et al., 2004), a relatively weak
correlation was found between KOH protein solubility and in vivo
characteristics which indicated that this index was not sufficiently
sensitive for evaluating protein quality of under-processed FFSB.
Correlation coefficients for UA were comparable with those for TIA
and PDI thus suggesting that, under conditions of the present study
(i.e. using FFSB not impaired by over-processing), this assay may
be a good predictor of chicken performance. However, as shown by
Dale et al. (1986) and Parsons et al. (1991), UA is unsuitable for
detecting over-processed FFSB or SBM samples. There was a
negative correlation between TIA and relative pancreas weight but,
in contrast to other reports (Herkelman et al., 1991, Perilla et al.,
1997, Clarke and Wiseman, 2005), only a moderately strong
relationship between the variables was observed. Even though
significant differences were found between most of the treatments,
pancreas weights in treatments T1 to T7 and T10 ranged within a
relative narrow limit, thus suggesting that the pancreas to BW ratio
might not be a sensitive indicator of TIA at lower trypsin inhibitor
levels. Nevertheless, linear regression model relating the present
growth performance data to relative pancreas weight showed a
good relationship between the variables. The regression equations
were y = 2532 - 286.4x (R2 = 0.78) and y = 1.301 + 0.1861x (R2 =
0.77) for WG and FCR, respectively.
Table 6 - Effects of soybean treatment on growth performance of chickens
Feed intake (g/d) Weight gain (g/chick) Feed conversion ratio
Treatment
1-10 d 11-24 d 25-35 d 1-35 d 1-10 d 11-24 d 25-35 d 1-35 d 1-10 d 11-24 d 25-35 d 1-35 d
a c d d c d a a a
T1 28.2 72.3 133.8 79.0 d 137 602 560 1299 d 2.07 1.68 2.66 2.13 a
a c cd c b cd b b ab
T2 28.1 76.1 142.6 83.2 cd 157 737 664 1558 c 1.79 1.45 2.37 1.87 b
a ab bcd ab a bc c b bc
T3 29.0 88.3 146.0 89.4 bc 211 875 756 1842 b 1.38 1.42 2.15 1.70 cd
a ab ab ab a abc c b bc
T4 29.3 91.0 162.0 95.6 ab 220 930 791 1940 ab 1.34 1.37 2.26 1.73 bcd
a b abc b ab abc c b bc
T5 27.9 85.4 153.5 90.4 b 204 837 782 1823 b 1.38 1.43 2.17 1.74 bc
a a a ab a ab c b bc
T6 29.5 93.5 166.5 98.0 a 228 953 880 2060 a 1.30 1.38 2.08 1.66 cd
a ab abc ab a abc c b bc
T7 29.8 91.6 158.1 94.8 ab 221 930 791 1942 ab 1.35 1.39 2.21 1.71 cd
a ab ab ab a a c b c
T8 29.0 91.8 161.1 95.5 ab 223 945 933 2101 a 1.30 1.36 1.91 1.59 cd
a ab ab ab a a c b c
T9 31.2 88.8 159.0 94.3 ab 229 939 917 2086 a 1.36 1.32 1.92 1.58 d
a ab bcd a a abc c b bc
T10 30.9 90.0 147.8 91.3 ab 232 882 800 1914 ab 1.33 1.44 2.06 1.68 cd
Pooled SEM 0.79 1.71 3.48 1.50 5.5 26.6 32.6 41.2 0.051 0.033 0.084 0.032
a.b.c.d
Means within a column not sharing a common superscript were significantly different (Tukey HSD test. P < 0.05).
19
Table 7 - Effects of soybean treatment on carcass characteristics

of chickens
Body weight Carcass Breast meat Pancreas
Treatment
(g) (% BW) (% BW) (g/kg BW)
e d c a
T1 1447 62.46 16.44 4.470
d c b b
T2 1677 64.92 18.43 2.920
bc abc ab f
T3 1972 66.54 19.87 1.850
ab ab ab e
T4 2115 67.08 19.33 2.000
c bc ab c
T5 1892 66.23 19.00 2.567
a a a g
T6 2183 67.94 20.25 1.734
abc ab ab d
T7 2046 66.94 19.31 2.258
a ab a f
T8 2197 67.00 20.17 1.848
a ab ab f
T9 2159 67.31 19.79 1.856
ab ab ab e
T10 2061 67.69 19.45 2.062
Pooled SEM 15.1 0.292 0.292 0.0222
a, b, c, d, e, f, g
Means within a column not sharing a common superscript were significantly
different (Tukey HSD test, P < 0.05).
Table 8 - Pearson correlation coefficients describing the relations

between in vivo chicken parameters and in vitro
indicators of FFSB treatment1
Feed Pancreas
Feed intake Weight gain
Item conversion weight
(g/d) (g)
ratio (g/kg BW)
KOH protein solubility (%) -0.67 *** -0.65 *** 0.49 *** 0.44 *
Urease activity (mg N/g) -0.81 *** -0.87 *** 0.78 *** 0.78 ***
Protein dispersibility index (%) -0.81 *** -0.88 *** 0.84 *** 0.85 ***
Trypsin inhibitor activity (mg/g) -0.85 *** -0.89 *** 0.79 *** 0.80 ***
1
Calculated for treatments T1 to T7
* P < 0.05; ***P < 0.001.
Digestibility trial
Producing a feed that meets the nutritional requirements of

the animal requires a precise knowledge about the nutrient and
energy content of the used feed ingredients. With regard to amino
acids, it is the most precise way to formulate feed on basis of
standardized ileal digestible amino acids which is only possible if
the necessary digestibility coefficients are known for the used feed
ingredients.
The total ileal AA outflow contains the non-digested dietary
AA and the endogenous losses. The endogenous losses represent
endogenously synthesized proteins and other AA containing
compounds that are secreted into the intestinal lumen of the
animal and are not digested and reabsorbed prior to end of the
ileum (e. g. digestive enzymes, mucoproteins, sloughed epithelial
cells, serum albumin, amides). The endogenous losses can be
separated into basal losses and specific losses. The basal
endogenous losses are not influenced by feed ingredient
composition. In contrast, the specific endogenous losses are
characteristic for different feed ingredients and are influenced by
their composition (anti-nutritional factors: trypsin inhibitors, lectins,
tannins; amount and structure of dietary fiber). Correcting the ileal
outflow for basal endogenous losses, makes SID AA values
additive in feed formulation (Stein et al., 2005) because the
calculation accounts then for any components that are specific to
the feed ingredient. If a standardized amount of basal endogenous
AA losses is used for correction, one yields the SID AA. The SID
AA are state of the art in diet formulation for pigs (Stein et al.,
2007) and poultry (Ravindran et al., 2017).
22
Table 9 - Average standardized ileal digestible amino acid coefficients (%) and 95%
confidence interval of full-fat soybeans determined in the actual trial
Trt. CP Met Cys M+C Lys Thr Trp Arg Ile Leu Val His Phe
T1 42.8 46.0 33.6 39.6 50.8 41.2 35.2 48.6 35.4 35.4 33.8 50.4 32.5
36.0-49.6 39.5-52.5 25.4-41.8 32.4-46.8 44.5-57.1 34.2-48.2 27.6-42.8 41.6-55.6 27.9-42.9 27.9-42.9 26.3-41.3 42.7-58.1 28.5-43.9
T2 57.9 55.8 39.2 47.3 63.0 52.3 47.0 65.0 48.2 50.8 49.0 62.3 53.5
54.4–61.7 52.7-59.0 34.3-44.1 43.5-51.1 60.2-65.8 48.6-56.0 42.6-51.4 63.1-68.0 44.0-52.4 46.9-54.7 45.0-53.0 59.2-6534 49.2-57.8
T3 69.2 74.8 59.3 66.7 73.5 68.3 69.7 72.7 67.0 68.2 67.0 74.7 68.2
63.8-74.6 70.1-79.6 53.3-65.4 61.3-72.0 68.5-78.5 63.2-73.5 64.5-74.9 67.5-77.8 60.9-73.1 62.3-74.1 61.2-72.8 69.5-79.8 61.9-74.4
T4 70.7 77.3 61.2 68.7 75.3 70.8 72.5 75.0 70.7 70.8 70.2 75.8 70.2
65.2-76.1 74.0-80.6 56.4-66.0 64.5-72.9 71.5-79.2 66.8-74.9 68.4-76.6 70.9-79.1 66.5-74.8 66.7-74.9 65.7-74.6 71.4-80.3 64.9-75.5
T5 75.0 77.5 60.8 68.7 79.2 72.0 70.5 80.7 72.0 72.7 71.5 79.2 74.3
69.6-80.4 71.9-83.2 52.8-68.9 62.0-75.3 74.5-83.8 65.9-78.1 63.7-77.3 76.2-85.2 65.9-78.1 67.1-78.2 65.3-77.7 74.3-84.0 68.7-80.0
T6 69.5 76.3 61.7 68.7 75.5 70.2 70.8 75.0 69.8 70.3 69.5 75.8 70.2
66.4-72.6 74.0-78.6 57.8-65.5 65.8-71.5 72.9-78.1 68.2-72.2 68.3-73.3 72.2-77.8 67.1-72.6 67.7-73.0 66.9-72.1 72.7-79.0 66.3-74.0
T7 76.0 81.2 63.8 71.8 80.3 74.8 76.7 79.8 76.8 77.0 76.3 81.3 77.0
74.2-77.8 79.7-82.6 61.4-66.3 70.2-73.5 79.1-81.5 73.1-76.5 75.0-78.3 78.2-81.5 75.1-78.5 75.1-78.9 74.5-78.1 79.5-83.2 74.6-79.4
Z1 80.8 85.3 67.8 76.2 84.2 79.7 81.5 85.2 81.5 81.8 81.2 85.2 81.3
79.3-82.4 83.8-86.8 64.6-71.1 74.1-78.3 82.9-85.5 77.9-81.5 80.1-82.9 82.9-86.5 79.8-83.2 80.3-83.4 79.5-82.9 83.8-86.5 79.3-83.3
Z2 81.8 86.9 67.5 77.0 84.7 79.8 82.7 86.2 84.1 83.7 83.3 85.8 83.6
80.1-83.5 85.3-88.4 62.9-72.0 74.0-80.1 83.1-86.3 77.4-82.2 81.0-84.5 84.6-87.7 82.2-85.9 81.9-85.5 81.5-85.1 84.2-87.4 81.4-85.7
Z3 79.2 85.3 62.7 73.8 82.5 77.3 80.3 85.0 82.2 82.5 81.5 83.8 82.0
76.9-81.5 83.5-87.2 57.8-67.5 70.6-77.1 80.4-84.6 74.4-80.3 77.9-82.7 83.0-87.0 80.2-84.2 80.8-84.2 79.4-83.6 82.5-85.2 80.9-83.1
Z4 80.0 85.0 65.3 75.2 82.3 78.5 81.0 85.5 82.7 82.7 81.8 84.2 82.3
79.0-81.0 84.3-85.7 62.7-68.0 73.6-76.7 81.7-83.0 77.5-79.5 80.1-81.9 84.4-86.6 81.7-83.6 81.7-83.6 80.9-82.8 82.9-85.5 80.7-84.0
Z5 78.2 85.5 58.2 72.2 78.3 77.5 80.5 85.7 83.0 84.3 82.2 82.3 83.8
75.7-80.6 83.0-88.0 53.4-63.0 68.8-75.6 75.6-81.0 74.3-80.7 77.6-83.4 83.2-88.1 80.6-85.4 82.3-86.4 79.5-84.8 81.1-83.5 82.2-85.5
Z6 74.0 82.3 52.3 68.3 71.3 73.8 77.7 82.0 80.3 81.8 79.5 77.5 81.0
72.1-75.9 80.6-84.1 49.0-55.6 66.4-70.3 69.7-73.0 72.3-75.4 75.9-79.4 80.5-83.5 78.6-82.1 79.9-83.8 77.7-81.3 75.1-79.9 78.9-83.1
Z7 66.8 74.7 45.7 61.3 62.2 65.7 69.2 75.8 72.7 74.8 71.8 70.7 75.5
63.6-71.1 70.7-78.6 41.1-50.3 57.1-65.6 58.4-65.9 61.4-70.0 65.0-73.4 72.8-78.9 68.7-76.6 71.1-78.6 67.7-75.9 68.4-72.9 73.2-77.8
Z8 63.7 71.0 40.0 56.8 56.5 61.2 64.7 72.8 69.0 71.2 67.8 65.3 71.8
59.0-68.3 67.1-74.9 32.3-47.7 51.4-62.2 51.2-61.8 56.4-65.9 60.8-68.5 68.6-77.1 65.2-72.9 67.9-74.4 63.8-71.9 60.0-70.7 68.0-75.7
Z9 51.8 60.0 24.7 44.3 39.8 50.2 53.3 63.0 57.7 60.7 56.8 53.7 61.5
49.1-54.5 57.1-63.0 20.0-29.4 40.8-47.9 36.7-42.9 47.2-53.1 50.8-55.9 60.1-65.9 54.7-60.7 58.2-63.1 54.0-59.7 50.0-57.3 58.8-64.2
- IV International Symposium on Nutritional Requirements of Poultry and Swine
Table 10 - Student t-test values for the standardized ileal digestibility coefficient of
Lysine. In case of T6 and T7, the Kolmogorov-Smirnov test p-value is
reported. Non-significant differences are marked in bold
T2 T3 T4 T5 T6 T7 Z1 Z2 Z3 Z4 Z5 Z6 Z7 Z8 Z9
T1 0.0154 0.0006 0.0004 0.0001 0.0086 0.0086 0.0004 0.0003 0.0003 0.0006 0.0004 0.0023 0.0205 0.2111 0.0232
T2 0.0073 0.0006 0.0003 0.0050 0.0050 0.0000 0.0000 0.0000 0.0000 0.0000 0.0009 0.7348 0.0670 0.0000
T3 0.5824 0.1342 0.8928 0.0310 0.0076 0.0058 0.0149 0.0176 0.1360 0.4507 0.0059 0.0010 0.0000
T4 0.2407 0.8928 0.0310 0.0051 0.0035 0.0130 0.0154 0.2427 0.1044 0.0007 0.0003 0.0000
T5 0.4413 0.4413 0.0890 0.0671 0.2385 0.2390 0.7682 0.0193 0.0003 0.0001 0.0000
T6 0.0050 0.0050 0.0050 0.0050 0.0050 0.4413 0.0310 0.0050 0.0050 0.0050
T7 0.0310 0.0310 0.4413 0.4413 0.4413 0.0050 0.0050 0.0050 0.0050
Z1 0.6196 0.2153 0.0392 0.0064 0.0000 0.0000 0.0001 0.0000
Z2 0.1313 0.0319 0.0040 0.0000 0.0000 0.0001 0.0000
Z3 0.8854 0.0393 0.0000 0.0000 0.0001 0.0000
Z4 0.0332 0.0000 0.0001 0.0002 0.0000
Z5 0.0023 0.0001 0.0001 0.0000
Z6 0.0035 0.0019 0.0000
Z7 0.1207 0.0000
Z8 0.0007
23
The determined coefficients of standardized ileal amino acid

digestibility are presented in Table 9 as averages together with
their respective 95% confidence interval. The confidence intervals
are larger for under- and over-processed samples than for
adequately processed samples of Z1 and Z2. For example, the
confidence interval of the digestibility coefficient of Lysine covers
12.7 points for K1 (raw soybeans) and 10.6 points for Z8 (over-
processed) but only 2.6 points for Z2 (treatment with highest
digestibility). Multivariate ANOVA gave a P-value smaller than
0.001, indicating that the processing had a significant impact on
the digestibility coefficients. Overall, lowest coefficients were
estimated in raw soybeans (T1) and digestibility improved with
increasing processing intensity. Highest coefficients were recorded
in Z1 and Z2 and digestibility was reduced with further processing.
As an example, a t-test was done to assess the difference of the
digestibility coefficients of Lysine per treatment and Erro! Fonte
de referência não encontrada. 10 shows the P-values. Under-
processsing as well as over-processing can lead to a very similar
amino acid digestibility. For example, the digestibility coefficients
of Lysine of the under-processed FFSB of T1 and T2 are
statistically not different (P > 0.05) from the over-processed FFSB
of Z8 and that of Z8 and Z7, respectively. In Erro! Fonte de
referência não encontrada.10, the non-significant differences are
marked in bold. With regard to the optimum digestibility coefficient
of Lysine it can be seen that the treatments Z1 vs Z2, Z1 vs Z3,
and Z2 vs Z3 are not different from each other (P > 0.05). The
respective digestibility coefficients of Lysine are 84.2, 84.7 and
82.5 for Z1, Z2, and Z3 and form a kind of plateau.
Figure 1 - Hierarchical clustering of the treatments based on their

similarity (y-axis). The colour represents the digestibility
coefficients per amino acid or crude protein.
In general, the digestibility coefficients of all amino acids of

Z1 and Z2 were in close agreement based on hierarchical
clustering (Figure 1). Thus, mean values were calculated and
compared with literature data (Table 11). A closer look on the
individual digestibility coefficients reveals that the coefficients of
Cys are much lower than those of the other amino acids. In, Cys is
a cluster for itself which is quite distinct from the others. For Cys a
digestibility coefficient of 67.7% was determined for Z1+Z2
whereas the other coefficients range from 79.8% for Thr up to
85.7% for Arg. The digestibility coefficient of Cys determined in
this trial is 9.3 points lower than that published by INRA (2002) for
extruded FFSB. However, it is well in line with data reported by
Ravindran et al. (2014) for SBM. The mean standardized ileal
digestibility coefficient of Cys determined in 55 SBM using growing
broilers was 66%.
The digestibility coefficient of Met determined in this
experiment is almost identical with that one published by INRA
(2002). However, the coefficients of the other amino acids are
lower than that published by INRA (2002). The INRA values are
not covered by the upper 95% confidence interval. The difference
is smallest for His with -1.5 points and largest for Phe and Arg with
-5.3 and -5.5 points, respectively. The coefficient for Met+Cys is
heavily impacted by the poor digestibility of Cys. These difference
might be partly caused by the different assays used to estimate
the AA digestibility, as the INRA values were derived with adult
roosters whereas growing broilers were used in the actual
experiment.
The impact of over-processing can be seen in treatments
Z3 to Z9. The digestibility of all amino acids decreases with
prolonged autoclaving. For example the digestibility of Lys is
reduced from 84.7% in Z2 down to 39.7% in Z9.
Table 11 - Comparison of the standardized ileal digestibility

coefficients (mean of Z1 and Z2) determined in the
actual trial in comparison to literature data
Treatment CP Met Cys M+C Lys Thr Trp Arg Ile Leu Val His Phe
Upper c.I.1 82.5 87.2 70.3 78.4 85.4 81.2 83.2 86.7 84.2 84.0 83.6 86.5 84.0
Z1+Z2 81.3 86.1 67.7 76.6 84.5 79.8 82.1 85.7 82.8 82.8 82.3 85.5 82.5
Lower c.I.² 80.2 85.0 65.0 74.8 83.5 78.3 81.0 84.6 81.4 81.5 80.9 84.5 80.9
INRA³ 86 77 81 88 85 - 91 87 87 86 87 88
1
Upper 95% confidence interval.
² Lower 95% confidence interval.
³INRA, 2002. Extruded FFSB, Rooster assay.
SUMMARY AND CONCLUSION
In the growth trial it was shown that today’s broilers are able
to achieve good performances if they are fed with adequately
processed FFSB. Even high inclusion levels of FFSB are tolerated
well and adequately processed FFSB can be used as the main
protein source in broiler diets. Under the tested conditions broiler
seem to be less sensitive to TIA than anticipated. However, under-
processing of FFSB needs to be avoided as it is performance
depressive.
For the digestibility trial it can be summarized that the
standardized ileal amino acid digestibility of FFSB is highly
dependent on the processing. Under- and over-processing results
in decreased amino acid digestibility. The best digestibility seen in
this trial is lower than previously reported. This is especially the
case for Cys and Met+Cys. Therefore, it is recommended to
recheck and update the digestibility coefficients which are
currently used in feed formulations.
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SIMULATION MODELS: USE OF ENERGY BY
POULTRY
Nilva Kazue Sakomura¹, Nayara Tavares Ferreira¹, Rob Mervyn Gous²
¹Faculdade De Medicina Veterinária e Zootecnia, Unesp Univ Estadual Paulista, Via Ac. Prof.
Paulo Donato Castellane, s/n, Jaboticabal, SP 14884-900, Brazil; ²University of KwaZulu-Natal,
Pietermaritzburg, South Africa
INTRODUCTION
The amount of energy required by poultry to maximize

growth or egg production has been the subject of a considerable
amount of research in the past, yet controversy still exists
regarding the role of energy in controlling food intake, and the
most appropriate unit of energy to use when describing energy
utilization in poultry.
The concept of energy partitioning for maintenance and lipid
retention in the body has been studied and described in the
literature for more than 30 years (Byerly, 1979; Sibbald, 1982;
Sibbald and Morse, 1984; Emmans, 1987; 1989). Mathematical
models that have been used to estimate energy requirement have
helped to understand the mechanisms of energy utilization in
poultry. Such models, applied to growing and egg producing
poultry, have enabled nutritionists to feed birds more accurately, to
achieve their genetic potential, reduce lipid deposition, minimize
nutritional problems in egg production and reduce feed cost.
Understanding the process of growth and energy
metabolism of birds and the factors that affect them is essential to
minimize the losses caused by metabolic diseases due to rapid
growth (broilers) and increased production (commercial hens and
broiler breeders). In addition to the requirement for energy it is
important to consider the nutrients essential for growth and
reproduction, with the objective of improving carcass
characteristics, favouring protein deposition, decreasing lipid
accumulation and reducing production costs (Sakomura et al.
2004).
Growth models assist in understanding the importance of

different factors that affect and control feed intake. The concepts
used in simulation models play a vital part in advancing our
understanding of the control of feed intake (Forbes, 2007). Some
of the recent simulation models are based on physiological and
metabolic principles for determining feed intake and nutritional
requirements, resulting in greater precision in defining feeding
programs. This presentation focuses on the principles on which
simulation models developed at Unesp-Jaboticabal, are based
which have assisted greatly in improving our understanding of
energy utilization in poultry.
DO BIRDS EAT TO SATISFY THEIR ENERGY REQUIREMENT?
Some researchers suggest that birds eat to satisfy their

energy requirements, which implies that energy is always first
limiting in all feeds (Leeson and Spratt, 1985, Summers 1985,
Lilburn et al., 1987, Silva, 2014). These authors and others
(Leeson, 2000; Veldkamp et al., 2005) based their theory on the
fact that feed intake is usually decreased when high-energy diets
are fed. Based on this theory, nutrient concentrations in the diet
are often expressed per unit of metabolizable energy (Gonzalez
and Pesti, 1983). This theory needs to be questioned given that
birds often become excessively fat when fed diets low in protein
(Gous et al., 1990), that laying hens and broilers overconsume
energy when placed on a high nutrient density diet (Morris, 1968;
Fisher and Wilson, 1974), and that body lipid content is reduced
when broilers are placed on a high protein feed (Gous et al.,
2012).
Many studies show that birds do not control feed intake
according to the energy concentration of the diet. Richards (2003)
observed that modern chickens selected for fast growth do not
regulate voluntary intake to achieve an energy requirement. He
suggested that this change in the ability to adjust consumption in
broiler chickens is the result of selection for growth that may have
altered hypothalamic mechanisms that regulate hunger and satiety

(postprandial and preprandial mechanisms), but it was shown long
before this publication that birds and animals adjust food intake
according to the level of the first-limiting nutrient in the feed
(Emmans, 1987).
Several papers in the literature describe experiments in
which both energy and protein levels influenced voluntary feed
intake. In one of these, Gous et al. (1987) reported a study in
which three levels of energy with four levels of amino acids per
energy level were evaluated, and found that all diets containing
low protein (amino acids), independently of the energy level,
resulted in a significantly greater reduction in feed consumption
than that brought about by differences in energy content. In
addition, on these low protein feeds they observed lower feed
intake on the low energy than on the high-energy diet.
Forbes (2007) reported that animals are sensitive to
quantities of nutrients and can make appropriate choices
according to the animal's requirement. Therefore, if the feed is
deficient or unbalanced in one or more essential nutrients the
animal is underfed and disabled, influencing feed intake. This
author also suggests that when an imbalanced feed is offered, first
the animal changes feed intake to reach the desired consumption
to meet the first limiting (energy or protein) and then modulates the
intake to optimize the balance between the nutrients. Balch and
Campling (1962), mentioned by Forbes (2007), after reviewing the
control of voluntary feed intake in non-ruminants, concluded that:
"...food intake is unlikely to be regulated by any single mechanism
and… oropharyngeal sensations, gastric contractions and
distension, changes in heat production and changes in the levels
of circulating metabolites, may severally be implicated".
Imbalances between nutrients are a special case when
investigating factors that influence food intake. According to
Harper et al. (1970) and Forbes (2007), the primary response to
an imbalanced feed is a decrease in food intake. If the imbalance
is severe there is a large decrease in feed intake and growth, but if
the imbalance is slight, there may be a compensatory increase in

feed intake, since the animal tries to maintain nutrient intake
(Emmans, 1987). For diets containing nutrients above the
"requirement" there is little or no effect on food intake until the
nutrient concentration is so high that it causes a drop-in
consumption due to the toxic effects of the excess (Figure 1)
(Forbes, 2007).
Figure. 1 - Generalized diagram of the effects of a nutrient in feed

on voluntary intake of excesses and deficiencies
(adapted from Forbes, 2007).
Wijtten et al. (2004) in a study with different levels of lysine

in the diet of broiler chickens (9.1 to 14.4 g digestible lysine/kg
feed) did not observe a significant effect on feed intake, although a
decrease in weight gain and feed efficiency was observed, and an
increase in lipid content in the carcass with a decrease in dietary
lysine concentration.
An increase in lipid deposition in the body when lower levels
of amino acids are fed has been observed in dose-response trials
with broiler chickens, pullets and laying hens at Unesp-Jaboticabal

(Venturini, 2012, Bendezu, 2012, Sato, 2012, Bonato et al., 2015,
Silva, 2012, Donato et al.,2014, Ferreira et al., 2016). In these
studies, diets were formulated using the dilution technique, with a
wide range of amino acid levels (Lysine, Methionine + Cystine,
Threonine and Valine) at a constant energy level. As an example
of these studies, data of body protein and lipid deposition from a
methionine+cystine assay with broiler chickens from 1 to 14, 14 to
28 and 28 to 42 d of age is presented in Table 1 (Donato et al.,
2014). The greater deposition of body lipid in chicks fed diets with
lower amino acid and protein and the same level of energy in the
diet gives clear proof that the birds are not adjusting food intake to
meet their energy requirements, but to meet the limiting amino
acid requirement. If they were, there would be no change in the
lipid content of the chicks on the different feeds. In addition, it is
possible to observe that feed intake follows the same trend as
reported by Forbes, 2007 and presented in Figure 1.
Table 1 - Body protein deposition (BPd; g/bird/day), body lipid

content (BLc; g/kg) and feed intake (FI, g/bird/day) male
broilers as a function of methionine + cysteine intake
(Met + Cys, mg/bird/d), in the initial (1 to 14 d), growth
(15 to 28 d) and final (29 to 42 d) stages of growth
Level 1 – 14 d 15 – 28 d 29 – 42 d
Met+ Met+ Met+
BPd BLc FI BPd BLc FI BPd BLc FI
Cys Cys Cys
1 60 1.59 142 23.6 197 4.75 174 86.99 427 9.02 185 205.1
2 124 3.25 128 34.4 314 7.34 146 96.42 646 13 163 216.4
3 171 3.92 118 36.4 411 8.75 122 96.65 796 15.9 138 204.8
4 214 4.57 102 37.0 498 10.4 104 95.04 945 17.9 119 197.3
5 248 5.02 60.8 35.2 537 9.87 83.8 87.29 1078 17.2 90.9 189.2
6 282 4.82 61.8 35.4 611 10.6 59 85.61 1213 18.2 101 183.8
7 302 4.70 42.4 33.4 661 10.9 48 81.19 1331 18.3 110 177.4
Morris and Njuru (1990) provided feeds with a range of

protein levels (167-251 g CP/kg) to growing broiler chickens and
laying pullets. They found a decrease in body lipid from 167 to 81
g/kg in the broilers and from 87 to 29 g/kg body weight in the
laying pullets as dietary protein increased. They demonstrated in

this trial that there is a limit to the amount of feed that birds are
capable of consuming when presented with a low protein feed,
thereby, ratifies the theory that birds are capable of overcoming a
nutrient deficiency by increasing intake. Buyse et al. (1992)
reported that broilers fed lower protein diets (150g crude protein/
kg diet) increased feed intake in an attempt to meet their protein
requirements. Contrary to these results, a reduction in feed intake
on severely deficient levels of protein was observed in Ross but
not in Cobb broilers by Kemp et al. (2005) and Berhe and Gous
(2005). Similarly, Shariatmadari and Forbes (1993) provided diets
with variation in protein content (65, 115, 172, 225 or 280 g CP/kg)
for broilers and laying hens and found that feed intake was not
affected by the variation in dietary protein, except for the lower
protein content, where food intake was severely reduced in both
strains. Furthermore, the carcasses of the birds that received diets
with 115 and 172 g/kg had a higher lipid content than those fed
higher protein diets.
A similar study was conducted at Unesp - Jaboticabal,
where two strains of commercial laying hens (ISA Brown and Hy-
line) were compared to evaluate the variation in balanced protein
levels (120, 140, 160, 180, 200 and 220 g of protein/kg feed). It
was observed that the birds fed deficient diets had higher feed
intakes and, in this way, they minimized the impact of the low
protein feeds. Because these birds tried to consume enough feed
to meet their protein requirements they overconsumed energy and
deposited this energy excess as body lipids (Bendezu et al. in
press).
Based on the studies mentioned, it becomes evident that
feed intake might seldom be influenced by dietary energy content
and that essential nutrients such as amino acids, minerals and
vitamins are more likely to result in overconsumption of energy if
these nutrients are not in sufficient concentrations in the feed. As a
result of this compensatory increase in feed intake caused by a
limiting nutrient there is an increase in body lipid deposition due to
the overconsumption of energy. In contrast, birds that are fed a

high protein diet may be able to reduce the intake of such a feed
by making use of body lipid as a source of energy, if excess of
energy is available in form of body lipid (Gous et al. 2012).
However, in cases where the dietary protein content is excessively
high, such as in pre-starter feeds for broilers and starter feeds for
turkeys, a point is reached where insufficient energy is available to
process the dietary protein and the efficiency of protein utilization
declines causing a reduction in growth and body lipid content
(Gous, personal communication). De Greef and Verstegen (1995)
reported that if energy intake is limited, as is sometimes the case
when pigs are control-fed, protein deposition will be less than the
maximum deposition potential; in addition, excess protein and
amino acid intake results in a higher energy requirement due to
energy expenditure for nitrogen excretion (Sklan and Plavnik,
2002).
ENERGY UNITS FOR FEEDS
Whereas many energy units have been proposed to

express the energy content of feeds for poultry (ME, AME, AMEn,
TME, TMEn and EE), the metabolizable energy (ME) system has
conventionally been used. Emmans (1984) has proposed an
alternative energy scale, the effective energy (EE), which
describes the amount of feed energy that is available to the bird
for maintenance and growth or egg production, and which is
equivalent to the net energy (NE) system generally used for pigs.
In this system, the heat increment (HI) associated with feed
consumption is a function of what the animal does with the feed
hence it is possible to calculate the amount of heat generated by
the bird when consuming this feed. The ME system does not allow
such calculations to be made.
Gous (2010) reports that there are many advantages in
using the EE system, since this system estimates with greater
accuracy than does the ME system the energy available for
maintenance and growth or egg production. The EE system

makes it possible to predict the heat production of a bird according
to its physiological and nutritional state and hence to determine
whether the bird is able to lose to the environment all the heat
necessary to enable it to achieve its desired food intake whilst
maintaining thermal equilibrium. Likewise, when the ambient
temperature is below the critical lower temperature for the bird, the
additional amount of energy required for it to remain in thermal
equilibrium can be determined. It has been thought in the past that
the adverse effects of high temperatures on performance could be
reduced through the use of highly digestible nutrients, such as
amino acids (Waldroup et al, 1976), or by replacing carbohydrates
with digestible fats thereby reducing the heat increment of the feed
(Dale and Fuller, 1979; 1980; Sinurat and Balnave, 1985).
However, Gous & Morris (2005) have shown that these strategies
are ineffective in reducing the heat increment sufficiently. Under
such circumstances, it is more useful to reduce the environmental
temperature in the poultry house.
In the Models developed at Unesp–Jaboticabal the EE
system of Emmans (1994) has been favoured because of its ability
to deal with issue of heat production in broilers and laying hens
when predicting food intake in these birds.
MODELS FOR ESTIMATING REQUIREMENTS
In the literature, there are many equations that are used to

calculate the energy requirement for poultry (Emmans, 1974;
Peguri and Coon, 1988; NRC, 1994; Sakomura et al., 1993, 2006).
The factorial model has been the basis for these models,
estimating requirements by considering differences in body weight,
body composition, potential for growth and reproduction of the
animals, and in some, accounting for the environmental
temperature. The factorial method is based on the principle that
the requirement for energy and for essential nutrients such as
amino acids is the quantity to be supplied to meet the
maintenance requirements and for the growth of body protein and

adipose tissue and/or for egg production (Sakomura and
Rostagno, 2016).
The energy requirement for maintenance (EMm) is related
to body weight, however, due to the variation in body composition
in lipid and water according to the age of the animals and the
manner in which they have been fed, Emmans (1987) suggests
that maintenance requirements should be expressed in terms of
body protein weight and the degree of maturity of the bird. The
factorial model to estimate the energy requirement still considers
the energy need for growth (Eq. 1), that is, all energy retained for
protein and lipid deposition is considered.
In the case of laying hens and broiler breeders, the energy
need for egg production is also computed, compartmentalized in
each component (albumen, yolk, and shell). The effective energy
requirement (EEr) for birds in production can be determined by the
equation (Emmans, 1994) (Eq.2):
Model for growing birds:

EEr=(1.63×𝑃m0.73×PBt)+(50RP+56RL) Eq.1
Model for laying hens:

EEr=(1.63×𝑃m0.73×PBt)+(50RP+56RL)+(0.025×YW+0.0036×AW+0.0012×SW) Eq.2
Where Pm body weight at maturity (kg); PBt is the body

protein weight at time t, RP and RL are the retentions of protein
and body lipid respectively. 1.63 MJ, which is the energy need per
unit of maintenance per day, 50 and 56 MJ are the amounts of
energy need for retention of protein and lipid (g/d), respectively.
0.025×YW, 0.0036×AW and 0.0012×SW are the amounts of
energy required to produce one gram of yolk, albumen and shell
(MJ/kg), respectively (Emmans and Fisher, 1986).
The same concept is used to calculate the requirement for
amino acids considering maintenance, growth (Eq.3) and egg
production in the case of laying hens and breeders (Eq.4).
Model for growing birds:

Eq.3
Model for poultry in production:
Eq.4
Where AAr is the digestible amino acid requirement (mg/d).

AABm is the amino acid requirement for the maintenance of body
protein (mg/PBm0.73 u / day), (Table 2), BPm0.73 is the body protein
weight at maturity (kg), and u is the degree of maturity
(u=BPt/BPm). BPt is the body's protein content at time t. FL is the
feather protein loss (g/d); FP is the protein weight of feathers at
time t, (g/d); AAp is the amino acid content in the feather protein
(mg/g). BPd is the rate of deposition of body protein (g/d). BPp is
the rate of deposition of protein in feathers (g/d). AABg is the
amino acid composition of the ideal protein (8 g/kg) for body
growth (Table 2) and k is the efficiency of use of dietary amino
acids for body and featherprotein deposition (Fisher, 1998). AAy is
the amino acid content in the protein of the yolk, CNg is the
nitrogen content in the yolk (27 mg N/g), AAa is the amino acid
content in albumen protein, CNa is the nitrogen content of
albumen (17 mg N/G) and k2 is the efficiency of utilization of the
amino acid deposition on the egg components.
Table 2 - Amino acid compositions (mg / g) used to estimate

amino acid requirements (de Lunven et al., 1973)
Growth Feathers* Maintenance Yolk Albumin
Arginine 68 65 50 434 330
Cystine 11 70 35 163 178
Histidine 26 8 10 148 132
Isoleucine 40 40 50 348 331
Leucine 71 70 32 548 521
Lysine 75 18 73 477 378
Methionine 25 6 25 175 240
Methionine + Cystine 36 76 60 338 418
Phenylalanine 40 45 16 261 368
Phenylalanine + Tyrosine 71 95 32 514 625
Tyrosine 31 50 16 253 257
Threonine 42 44 40 313 272
Tryptophan 10 7 10 121 116
Valine 44 60 60 378 429
*Emmans (1989).
These models are based on the theory integrated by

Emmans and collaborators (Emmans, 1981; Emmans & Fisher,
1986; Emmans & Oldham, 1988; Emmans, 1989; Martin et al.,
1994) that estimate the amino acid requirements for growing
animals from based on the growth and body composition of an
individual of a given genotype. It is assumed that commercial
laying hens cease to grow body and feather protein once they start
producing eggs, and in that case the part related to body protein
growth, (50RP) and , is not used in the equation to
calculate the energy and amino acid requirements (Eqs. 2 and 4),
respectively. Body lipid might still be deposited in hens when a
feed deficient in an essential nutrient is fed, which results in the
hen overconsuming energy; and body lipid may be used as an
energy source where possible.
DESIRED FEED INTAKE (DFI)
Every animal has a purpose, and that is to grow as fast as

possible to reach sexual maturity as soon as possible so that it
can pass on its genes to the next generation (Emmans, 1987).
This implies that an animal will attempt to consume sufficient of a
given food to achieve this objective. By definition, the DFI is the
amount of food the bird will attempt to consume to meet the
requirement for the first limiting nutrient in that feed (Emmans and
Fisher, 1986) and this is calculated by dividing the requirement for
each essential nutrient by the dietary content of that nutrient
thereby identifying the DFI for the most limiting nutrient in the feed.
The daily requirements are calculated using equations 1 and 3 for
growing birds and 2 and 4 for laying hens. The DFI based on the
energy requirement and feed content is also calculated, the unit of
energy used being the effective energy (EE) (Emmans, 1994).
As an example of how the DFI for broilers is calculated the
requirements for EE, lysine and methionine + cysteine (M+C) are
estimated based on the data in Table 3, and feeds with different
concentrations of these nutrients (Table 4).
With these data, the requirements for energy, lysine and
M+C are calculated, according to the equations below:
0.73
EEr = (1.63×0.9 ×0.27) +(50×0.0171+56 ×0.0081) = 1.646 MJ/ bird d
0.73
AALys=(8×73×0.9 ×(0.27/0.9))+(0.01×52.8×18)+((16.2×75)/0.6)+((18×0.176)/0.6)=
164 mg lysine/bird d
0.73
AAM+C=(8×60×0.9 ×(0.27/0.9))+(0.01×52.8×76)+((16.2×36)/0.6)+((76×0.176)/0.6)
=134 mg M+C/bird d
In this example the EE of the feed was assumed to be 0.88

of the ME. Table 4 presents some examples of dietary energy,
lysine and M+C concentrations, the DFI calculated as a function of
the calculated requirements, and the limiting nutrient defined by
the highest DFI.
Table 3 - Data used to estimate the energy, lysine, and

methionine+cysteine (M+C) requirements for broilers
Data
Body weight (BW), kg 1.50
Body protein content, g/kg BW 180
Weight of feathers, g/kg BW 40
Body lipid content, g/kg BW 85
Body protein at maturity, kg 900
Gain in body weight, g 90
Feather loss, g/d 0.2
Calculations
Body protein weight (PBt) = 1.50x0.180 = 0.27 kg
Deposition of protein in the body (RP) = 90x0.180= 16.2 g/d
Feather weight = 1.500x40/1.0 = 60 g
Protein in feathers (PF) ¹ = 60x0.88 = 52.8 g
Lipid deposition in the body (RL) = 90x0.085 = 7.65 g/d
Feather protein deposition (DBPp) = 0.2x0.88 = 0.176 g/d
Efficiency of utilization of the amino acid for deposition in the body and feathers
(k) = 0.6
¹Assuming 0.88 g of protein/g in feathers.
Table 4 - Desire feed intake (DFI) based on dietary nutrient

concentrations and requirements for energy (1.646 MJ/
bird d), lysine (164 mg/bird d) and methionine +
cysteine (M+C) (134 mg/bird d)
DFI DFI DFI
ME EE Lysine M+C First
EE Lys M+C
limiting
kcal/kg MJ/kg g/kg g/kg g/d g/d g/d
12.3 10.3 134 130 Energy
2960 10.90 11.3 9.3 151 145 144 Energy
10.3 8.3 159 162 M+C
12.3 10.3 134 130 Energy
3000 11.05 11.3 9.3 149 145 144 Energy
10.3 8.3 159 162 M+C
12.3 10.3 134 130 Energy
3250 11.97 11.3 9.3 137 145 144 Lysine
10.3 8.3 159 162 M+C
The example given in Table 4 illustrates the point that for a

given animal on a given day small differences in the concentration
of dietary nutrients will change the order of the limiting nutrients
thereby altering the first-limiting nutrient and the DFI.
Of course, it is not always possible for the bird to
compensate completely for very low levels of nutrients because
feed intake may be constrained by the bulkiness of the feed
(insufficient gut capacity to deal with the bulk needed) or, more
often, the bird cannot lose to the environment the amount of heat
generated by the excessive amount of food that would be needed.
Because the bird must retain a constant body temperature it
cannot increase food intake indefinitely, and the external
environmental conditions will dictate how much the bird can
consume without increasing body temperature. When food intake
is constrained the bird will be unable to grow or reproduce at its
potential.
EFFECT OF UNDER AND OVER-FEEDING PROTEIN ON THE

BODY COMPOSITION OF BROILER CHICKENS
Because food intake increases as broilers grow the

concentration of dietary protein required is reduced, but it is
impractical to reduce the dietary concentration every day, hence
the use of a phase-feeding programme. In commercial broiler
production, feeding programmes are usually divided into phases,
using three to five diets decreasing in protein throughout the
production cycle. Assuming that the birds are given ad libitum
access to feed they are therefore generally provided with sub-
optimal levels of protein (amino acids) at the beginning of each
phase and excesses towards the end of each phase as illustrated
in Figure 2.
At the start of each phase the birds will attempt to consume
sufficient of the limiting amino acid(s) by overconsuming energy
which is deposited as body lipid. Feed efficiency is poor during
this period. As each phase progresses the under-supply of amino
acids becomes less and then at a point will become excessive in

relation to the requirement. At this point the bird is able to draw on
the excessive body lipid supplies to obtain much of the energy
required, and feed efficiency is improved.
Figure 2 - Dietary protein content supplied to birds in a three-

phase feeding systemin relation to their daily
requirement for protein (Buteri et al., 2009).
In order to elucidate the effect of under- and over-feeding

protein on body composition of broilers AVINESP software was
used to perform simulations of different nutritional programmes in
relation to body composition.
This paper does not attempt to describe the Avinesp
software since a full description can be found in Hauschild et al.
(2015). The simulations presented here were performed using the
Avinesp Model to demonstrate the possibility of using models as a
diagnostic tool, enabling nutritionists to improve the efficiency of
feeding broilers.
Six feeding programmes were simulated, divided into three
phases: Initial (1 to 21 d), Growth (22 to 35 d) and Finisher (35 to
42 d), two energy levels with three levels of protein for each phase
(Table 5).The DFI’s for each feeding programme simulated are
shown in Figure 3. Feed composition directly influences DFI since
the bird attempts to consume sufficient feed to meet the

requirement for the first limiting nutrient. Thus, diets with low
protein (program 3–HE/LP and 6–LE/LP) result in a higher feed
intake (DFI) and ME intake (Figure 3), consequently, higher lipid
deposition in the body (Figure 4). In addition, on the lower protein
feed the bird cannot consume sufficient lysine to meet the
requirement (Figure 5) with the result that there is also a reduction
in body protein deposition (Figure 6). Gouset al. (1990) report that
birds fed unbalanced diets, where energy is not first limiting, retain
lipid in excess.
Table 5 - Feeding programme as a function of the age (d) and

energy (kcal/kg) and protein (%) level used in each
simulation
Programmes Starter (1 to 21 d) Growth (22 to 35 d) Finish (35 to 42 d)
1 – HE/HP 3250 ME / 26 CP 3350 ME / 24 CP 3400 ME / 22 CP
2 – HE/MP 3250 ME / 22 CP 3350 ME / 20 CP 3400 ME / 18 CP
3 – HE/LP 3250 ME / 18 CP 3350 ME / 16 CP 3400 ME / 14 CP
4 – LE/HP 3050 ME / 26 CP 3150 ME / 24 CP 3200 ME / 22 CP
5 – LE/MP 3050 ME / 22 CP 3150 ME / 20 CP 3200 ME / 18 CP
6 – LE/LP 3050 ME / 18 CP 3150 ME / 16 CP 3200 ME / 14 CP
Figure 3 - Desired feed intake as a function of age for each feed

program simulated.
Programs 4–LE/HP and 5–LE/MP (lower ME and higher

and medium CP) provided ME intakes close to requirement (Fig.
4) and lipid deposition according to genetic potential. The program
5–LE/MP (lower ME and medium CP provided ME and lysine
according to requirements (Fig.’s4 and 5), consequently higher
protein deposition and lipid deposition according to genetic
potential (Fig 4). These results show that when ME/CP is suitable,
lipid deposition in the body is according to genetic potential.
The effect of under- and overfeeding on each feeding phase
(Figure 2) may affect body composition, as observed in program 1–
HE/HP, 2–HE/MP and 5–LE/MP. There is an increase in energy
intake, especially when changing the diet phase, and as a
consequence there is a greater deposition of body lipid (Figure 4).
Emmans (1981) reported that body lipid reserves are labile, changing
on account of feed level, previous feed offered, and environmental
conditions. In addition, growing chickens with ad libitum access to feed
with low first-limiting amino acids to energy ratio take more time and
eat more feed energy to reach a given weight, and are also fatter at
that weight (Jackson et al., 1982; Gous et al., 1990).
50
Figure 5 - Lysine requirement (dotted line), lysine concentration in the feed (dashed line) and lysine intake corrected
by the AFI (line) according to feeding program.
Figure 6 - Body protein content (g) and body lipid content (g) of
broilers in different feed programs as a function of age.
The feed intake in dependent, among other factors, on the

state of the animal at the time, and this feed intake will define the
amount of protein and lipid that will be deposited each day (Gous
et al., 2012). In Figure 6 there is a small variation in the deposition
of body protein among the 1–HE/HP, 2–HE/MP, 4–LE/HP and 5–
LE/MP programs, except for programs 3–HE/LP and 6–LE/LP,
because, of the low concentration of protein in the diet at all
stages, the body protein deposition was lower than observed in
other programs. In addition, program 3–HE/LP showed the highest
lipid deposition, since the birds increased feed intake to reach the
lysine requirement, and programs1–HE/HP, 4–LE/HP and 5–
LE/MPhad the lowest lipid deposition, since the diet provided
sufficient lysine and energy.
EFFECT OF ENERGY INTAKE AND LIPID DEPOSITION OF

LAYING HENS AND BROILER BREEDERS.
Simulation of the effect of dietary energy and lysine on egg

production in laying hens:
Laying pullets and broiler breeders do not reach somatic

maturity by the time they become sexually mature and start laying
eggs. At the time that egg production commences pullets cease
depositing body and feather protein, so there is no obligatory
requirement for body and feather protein growth in these birds
(Gous & Nonis, 2010). Changes in body weight during the laying
period will therefore be the consequence of the way birds are fed,
low protein feeds, for example, leading to greater deposition of
lipid. Thus, an important factor to be considered in the nutrition of
laying hens is the use of dietary energy in these birds. There are
three main requirements for dietary energy: maintenance,
production (lean tissue or egg production) and storage as lipid.
To better understand the mechanism of dietary energy
utilization and the mobilization and deposition of body lipid in
commercial laying hens, a simulation was done using the
simulation model developed at Unesp Jaboticabal. A population of
laying hens from 18 to 60 weeks of age was simulated considering
the production potential according to the Hy-Line breeder manual
and the responses according to the simulated diets. This model
considers the requirements of energy and amino acids for
maintenance and egg production calculated by the equations
presented in item 1.4 (Eq. 1 and 2). In this simulation three energy
levels (kcal of ME/kg) and three lysine levels (%) were applied,
namely, 2600/0.8; 2600/0.7; 2600/0.6; 2700/0.8; 2700/0.7;
2700/0.6; 2800/0.8; 2800/0.7 and 2800/0.6. Figure 7 shows the
mobilization and deposition of body lipid, the DFI to meet the
requirement of the first limiting in the diet, and the rate of laying in
response to dietary energy and lysine levels.
In the feeding programmes with low lysine (0.6%) feed intake is

increased to the same extent at all energy levels and consequently a
higher body lipid deposition was observed in response to the low
lysine inclusion in the diet, being higher in (2700 and 2800). At 0.7%
and 0.8 % of Lys the ME levels also influences feed intake at peak
with intake decreasing as the ME is increased (115, 110 and 105 g).
However, inclusions of 0.7% and 0.8% lysine were not limiting, since
there was no difference in feed intake at the same energy level. The
higher level of dietary energy reduced feed intake, and the increase in
lysine did not alter DFI, a slight trend has been observed for increasing
consumption from the highest energy level to the lowest. This results
show that when lysine is limit (0.6) the birds increase the feed intake,
in higher levels (0.7 and 0.8) the feed intake does not change, in these
levels the ME were limit, since birds increased the feed intake as ME
levels decreased. This trend is in line with that observed by
Murugesan and Persia (2013) who reported an increase in the intake
of 2 and 3 g with a reduction of 35 and 154 kcal ME/ kg, respectively.
Differences in body lipid content (Figure 7) illustrate well the
effect of feeding different ratios of lysine and ME to laying hens. Birds
increase food intake until peak. However, if they are fed adequate
levels of lysine they will mobilize body lipid and keep the increase in
food intake to a minimum whilst maintaining egg production. At the
lowest level of lysine, because of the high food intake, considerable
amounts of body lipid were deposited irrespective of the energy
content of the feed. As the dietary lysine content was increased lipid
deposition was reduced almost completely on the low energy feed, but
continued to increase to different extents on the higher energy feeds.
These results concur with those of Persia (2016) who reported a
significantly reduced abdominal lipid weight in response to decreased
dietary energy, with no change in egg production. This author
highlighted the chicken's ability to continue egg production by reducing
body lipid reserves to compensate for the reduction in dietary energy
content, at least in the short term.
Bobeck et al. (2014) also reported that lower dietary energy
resulted in a reduction in body lipid before egg production was
negatively affected, in spite of an increased intake, which was also

observed in this simulation (Fig. 7). Despite differences in energy
and lysine levels, egg production was not affected, except at the
inclusion of 0.6% lysine, which showed a drop-in egg production
from the 45th week of age (Fig. 7).
Figure 7 - Desired feed intake; deposition and mobilization of body

lipid (g); and rate of laying in response to energy levels
(2600, 2700 and 2800 kcal/kg) and lysine (0.8, 0.7 and
0.6%) in the diets of laying hens from 18 to 60 weeks of age.
Thus, the birds mobilize energy from lipid reserves to

achieve the genetic potential of egg production (Fig 7). It can be
inferred that laying hens prioritize egg production in relation to
body lipid storage, since it seems that body composition is a more
sensitive indicator of short-term dietary energy content than egg
production (Persia, 2016). Similarly, Emmans and Fisher (1986)
reported that hens will support egg production until the available lipid
reserves cease, and the time the birds will remain in production will
depend on how deficient the diet is and the lipid reserve available.
On the other hand, on days when birds do not produce
eggs, the energy requirement is lower relative to the day of
production. If the amount of energy consumed is greater than the
requirement for maintenance, the excess energy consumed will be
deposited, leading to a reserve of lipid on that day. Mobilization of
lipid reserves when needed, and the deposition of excess energy
as body lipid is a dynamic process and is only really observable
with the use of simulation models.
Simulation of the effect of feeding programmes on the use of

energy by broiler breeders
Broiler breeders, due to feed control supply, cannot increase

voluntary feed intake to obtain sufficient of a limiting dietary nutrient.
To better understand how lipid deposition and egg production are
affected by changes in the supply of amino acids and energy in broiler
breeders a stochastic model developed at Unesp-Jaboticabal was
used.
The model considers the requirements for energy and amino
acids for maintenance and egg production, calculated using the
equations presented in item 1.4 (Eq. 2 and 4). In this model the
genetic potential of the broiler breeders is input and the nutritional
requirements and consequent performance of the flock can be
simulated.
Laying performance of 200 broiler breeders, from 20 to 60
weeks of age, was simulated to predict the response of these birds to
six feeding programmes. For all feeding programmes, a diet with the
same composition (protein and amino acids) except for energy was
considered. The nutrient composition of the diets is given in Table 6.
The reproductive performance of the simulated population
(YW, AW, SW, EW, rate of laying, egg output and EE r) is
summarised at four-weekly intervals in Table 8. The maximum rate
of lay (83%) was achieved around 28 weeks of age and declined
after this to 49% at 60 weeks. During that time, mean EW

increased from 55 to 76 g, YW and AW increased from 12.7 to 23
and 37.2 to 46.6 g, respectively. The requirement for EE at four-
weekly intervals range from 1.09 to 1.36 MJ/ bird d, with the
largest requirement matching the highest egg production.
Table 6 - Diet composition (g/kg) in function of age (weeks)

20 to 24 w 25 to 36 w 37 to 60 w
Protein 150 160 150
Arginine 9.4 9.2 8.9
Isoleucine 6.2 6.5 6.4
Leucine 13.2 13.3 13.0
Lysine 6.3 6.7 6.6
Methionine 3.8 3.8 3.1
Methionine +Cystine 6.2 6.8 6.7
Threonine 5.0 5.0 4.9
Tryptophan 1.7 1.6 1.6
Valine 5.5 6.4 6.1
Table 7 presents the different feeding programmes used in

the simulations, considering a quantitative and qualitative
restriction or the combination of both.
Table 7 - Feeding programme, in function of the age (weeks) and

energy level (Kcal/kg) used in each simulation
Feeding programme 20-24 w 25-36w 37-60w Energy
Low feed intake and low energy (1-LFI-LE) 0.130 0.160 0.150
2800
High feed intake and low energy (2-HFI-LE) 0.150 0.170 0.160
Low feed intake and medium energy (3-LFI-ME) 0.130 0.160 0.150
2860
High feed intake and medium energy (4-HFI-ME) 0.150 0.170 0.160
Low feed intake and high energy (5-LFI-HE) 0.130 0.160 0.150
3000
High feed intake and high energy (6-HFI-HE) 0.150 0.170 0.160
From the results of the simulation of the population shown in

Table 8 the requirements for energy and amino acids for maintenance,
growth (to sexual maturity) and egg production of birds from 20 to 60
weeks of age, and then the DFI was calculated.
Table 8 - The mean body weight (BW), rate of lay (%), weight of yolk,
albumen and shell (g), and egg output (g/d) at four-weekly
intervals for simulated population of broiler breeders
Age BW %Lay Yolk Albumen Shell Egg weight Egg output
24 2.59 29.4 12.7 37.2 5.2 55.1 16.2
28 2.79 83.3 15.4 40.0 5.6 61.0 50.8
32 2.82 81.1 17.2 41.8 5.9 64.9 52.6
36 2.84 76.1 18.7 43.1 6.1 67.9 51.7
40 2.86 71.4 19.8 44.0 6.2 70.0 50.0
44 2.88 66.4 20.7 44.8 6.3 71.9 47.7
48 2.90 62.6 21.3 45.3 6.4 73.0 45.7
52 2.92 54.9 22.0 45.8 6.5 74.4 40.8
56 2.94 49.8 22.5 46.2 6.5 75.3 37.5
60 2.96 49.4 23.0 46.6 6.6 76.2 37.7
The EE, LYS and MET+CYS requirements and DFI of

broiler breeders from 20 to 60 weeks of age according to feeding
programmes are presented in Table 9.
Table 9 - The energy (EEr), LYS and MET+CYS requirements

(±SD) and DFI (g/bird/d) for broiler breeders of 20 to 60
weeks of age according to feeding programs
DFI
Age EEr LYS MET+CYS
P1 P2 P3 P4 P5 P6
week MJ/bird/d mg/bird/d mg/bird/d Kg
20 0.740±0.015 46.97±0.64 38.82±0.51 74.6 73.9 73.3 73.1 68.6 69.5
24 1.174±0.042 74.25±5.67 63.20±5.42 118.6 115.9 120.6 114.8 111.3 112.4
28 1.315±0.013 105.59±1.91 104.30±2.00 131.6 131.8 128.3 129.3 124.9 124.7
32 1.327±0.018 106.14±2.60 104.67±2.73 133.1 133.2 129.8 130.9 125.8 125.6
36 1.359±0.019 108.8±2.62 107.06±2.73 136.0 136.5 133.0 134.3 128.7 128.7
40 1.385±0.017 112.95±2.35 111.36±2.45 138.3 139.0 135.7 137.0 130.1 129.8
44 1.392±0.018 112.67±2.52 110.64±2.62 138.7 139.8 136.5 137.4 130.8 130.8
48 1.389±0.022 111.00±3.04 108.57±3.16 139.2 139.3 136.5 136.5 130.5 130.3
52 1.400±0.022 111.24±3.08 108.64±3.19 140.2 139.5 137.3 138.5 131.5 131.7
56 1.392±0.020 108.68±2.76 106.11±2.86 139.1 139.8 136.9 137.4 130.5 130.4
60 1.363±0.017 103.55±2.26 100.56±2.34 136.1 136.9 133.6 134.6 127.8 128.2
* P1 = 1-LFI-LE, P2 = 2-HFI-LE, P3 = 3-LFI-ME, P4 = 4-HFI-ME, P5 = 5-LFI-HE and P6 =6-HFI-HE.
It is observed an increase in the energy requirement from

0.7 to 1.3 MJ/day, from 20 to 25 weeks of age, because this period
coincides with the maturation of reproductive organs, this same
trend occurs for amino acids. Furthermore, it is possible to
observe that the DFI is higher than energy requirement for the
feeding programme 5-LFI-HE and 6-HFI-HE; due the high
concentration of energy in both programmes, which may be
inferred that DFI is not a function of energy (Fig.8).
Figure 8 - LYS, MET+CYS and EE requirements of broiler

breeders from 20 to 60 weeks of age and the DFI
according to feeding programmes.
The energy balance was calculated by difference between

energy intake and requirement (Energy Balance = EEintake -
EErequirement). As the model calculates the requirement for EE,
dietary ME was converted into effective energy (0.85ME). The
energy excess was converted into lipid deposition (56 MJ/kg lipid
retained) and the energy available for lipid mobilization was 39.6
MJ/kg lipid mobilized (Emmans and Fisher, 1986) (Fig. 9).
Figure9 - Body lipid content, body weight, egg production (%) and
egg weight according to feeding programmes, and egg
production potential of broiler breeders (dashed line).
In Figure 9 it is observed that even though there is an

increase in dietary energy content (3-LFI-ME), due to the higher
feed restriction body lipid deposition is lower than a programme
with high feed intake but with a lower energy content (2-HFI-LE),
because the bird cannot meet the requirement by regulating
voluntary feed intake. In addition, the birds fed 1-LFI-LE and 3-LFI-
ME start mobilization of body lipid reserves after 40 weeks but
cannot maintain egg production and egg weight (Figure 9). This
mobilization is described by Nonis and Gous (2012), who reported
that if feed intake is severely restricted lipid stores are
accumulated on days where the birds do not produce eggs to be
available when they are needed. However, birds are unable to use
all their lipid reserves when feed intake is severely restricted since
a minimum amount of body lipid must be retained in the body
(Wellock et al., 2003).
Nonis and Gous (2012) suggested that the minimum lipid

content should be about 110 g/kg body weight, which is higher
than that found in feeding programmes 1-LFI-LE and 3-LFI-ME.
Furthermore, it is possible to observe that in all feeding
programmes body lipid was mobilized from 20 to 23 weeks of age
due to the increased requirement to support the growth of the
reproductive organs and the onset of egg production. According to
Bowmaker and Gous (1989) the reproductive organs should be
considered in models to estimate the nutritional requirements,
since they suggest that the greater supply of energy and protein
can stimulate this development.
Silva (2014) reported that the pre-production diet (20 to 24
weeks of age) with higher ME content will allow a smoother energy
transition in the production phase, promoting adequate weight gain
and lipid deposition. Figure 9shows the rates of egg production
and egg weight according to feeding programmes. The feeding
programmes1-LFI-LE and 3-LFI-ME presented a decrease in egg
production and in egg size in compared to the genetic production
potential, because as discussed in item 1.5, the birds prioritize the
production of eggs by mobilizing body lipid, which can be
observed in Figure 8. However, there is a physiological limit to the
amount that can be mobilized and reproductive performance will
depend on the extent of the reserve of body lipid. Due to the
quantitative and qualitative restriction imposed by these feeding
programmes (1-LFI-LE and 3-LFI-ME) egg production could not be
sustained, and this was reduced below the potential after 42
weeks of age.
It is essential to provide the correct amount of energy to
meet the requirements for maintenance, support growth in the
period prior to the commencement of egg production and egg
production itself, mainly because broiler breeders are fed using a
controlled feeding programme. Based on energy utilization,
maintenance has priority, followed by growth of the yolk and
albumen. Variation in maintenance requirement maybe attributed
to differences in body weight and composition, and this can be
affected by genetics, feeds and feeding programmes and

management, and may affect lipid and protein deposition in body
tissues, lipid metabolism, egg composition, the size and metabolic
rate of the liver, gut, and reproductive tract (Renema et al., 2007).
Besides, egg output is dependent on energy intake when the
amino acid requirements for maintenance and growth of the yolk
are met. If the intake of the limiting nutrient on one day cannot
sustain this minimum egg size, egg production does not occur
(Gous and Nonis, 2010). The average chicken egg yolk contains
4 g of triglycerides, thus to support egg production a large lipid
(triglycerides) supply is required to support the demands of new
yolk formation (Richards et al., 2003).
CONCLUSION
Simulation models can be used by the nutritionist to define

feeds and feeding programs that will optimise performance and
result in higher profits for the producer.
The ratio of ME:AAs was shown by simulation to affect body
composition dynamically, thus the nutritionist can make decisions
about nutritional levels for broilers aimed at maximising the quality
of the carcass.
For laying hens and broiler breeders, simulation models can
assist in understanding how birds deposit or mobilize body lipid to
maintain egg production. They can help to establish nutritional
programs (nutrient levels and diet supply) to meet the
requirements of the bird to achieve good performance and
maximise profit. These results support the view that a modelling
approach has the potential to help nutritionists to make the right
decisions when feeding poultry.
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AMINO ACIDS IN BROILER NUTRITION: ALIGNING
PRACTICE WITH SCIENCE
Gabriel B. S. Pessoa, Eduardo T. Nogueira, Marianne Kutschenko,
Luciana F. de Lima
Ajinomoto do Brasil Ind. e Com. de Alimentos Ltda.
Rua Vergueiro, 1737, Vila Mariana, São Paulo, SP – 04101-000
ABSTRACT
Production of protein from animal sources will have to

increase more than 50% to help feed the world population in 2050
year. Moreover, that challenge become even harder when we
consider the limited land available for crops and animals. The
continuous investments and studies related to the broiler
production, by private companies and universities, are utmost
important to supply the world’s meat demand in the future. One
technology that have proven its efficiency is the use of the ideal
protein concept in the feed formulation together with the industrial
amino acids. This allows a) attending more precisely the animal’s
requirements on amino acids; b) to avoid excess of protein in the
diet, lowering the excretion of nitrogen to the environment; c) to
save arable land; d) to reduce the feed cost. The use of feed
supplemented with industrial AAs results in positives effects on
performance, economic and environmental parameters related to
the broiler production.
INTRODUCTION
According to FAO estimative, by the year 2050 the world

population will reach 9 billion people and the food production will
have to increase over 50% to attend the demand. From the animal
protein sources, it’s expected that broiler industry will have the
higher growth rate, achieving roughly 214 million tonnes of meat
produced by 2050. The continuous investments and development
in technology by industries and the evolution in the studies related
to animal nutrition and production are utmost important to

guarantee that the broiler industry will fulfil it important paper in
feeding the world.
To attend the need of food for the growing population, it
was needed to domesticate and raise the animals, nowadays most
of them in confinements, what generated some problems related
to the nutrition of these animals. Those facts aligned to the high
productivity aimed by the producers represented great stress to
the animals, creating the necessity of revision by the researchers
of the animal’s nutritional requirements. From that point, the
nutrition became a science.
This science started with the chemistries studying the
feedstuff composition. The feedstuffs are not complete in all
nutrients, and most of them are no well-balanced considering the
nutrients they have. This fact led to the conclusion that is
recommended to mix feedstuffs in order to achieve a well balance
among the nutrients. Further, the physiologists became interested
to know how the body works. The development in this area
allowed stablishing the needs of the organisms in terms of
nutrients, and subsequent fine tune led to the actual requirements
we know.
For many years, monogastric feed has been formulated
under the concept of crude protein (CP), often resulting in diets
containing amino acid levels higher than those do required. In the
body, carbon skeletons of excess amino acids (AAs) are used for
energy production, with residual nitrogen (N) being excreted by the
kidneys with a high-energy expenditure for the body.
With advances in scientific research in the area of nutrition
and animal metabolism, Harold H. Mitchell proposed the new
concept of ideal protein (Mitchell, 1964), which refers to a situation
where all essential AA are co-limiting for performance so that the
AA supply exactly matches the AA requirement. Together with
advances in the production technology of industrial AAs and at
compatible prices, it became possible to formulate diets with lower
protein levels without compromising the animal’s performance.
HISTORY OF AMINO ACIDS
The discovery of amino acids dates of the beginning of 19th

century. The first AA was described in 1806 by Vauquelin and
Robiquet when a quantity of juice from asparagus plant, which had
been concentrated by evaporation, was permitted to stand for
some time. Both chemistries described that this substance (the
asparagus juice) left no ash on ignition and that, on treatment with
nitric acid, it was decomposed with the liberation of nitrogen. This
substance was a puzzle to them and they concluded the
publication citing that the substance contained hydrogen, oxygen,
and carbon in definite proportions and probably nitrogen, but did
not give a name to the new product. Only in 1826, when
describing this work, Dulong refers to the substance as “which
they have designated as asparagine".
Vickery & Schmidt (1931) reported twenty-one amino acids
discovered until the year 1922 (Table 1). The amino acids are
arranged on the left side of table 1 in the order of their discovery,
either in nature or as the result of synthesis; the discoverer and
date are also given. On the right side of the table, the amino acids
are arranged in the order of their discovery as products of the
hydrolysis of proteins.
From this list, eighteen are among the common
proteinogenic amino acids currently known. Curiously, one the
most important AA in usage nowadays, the threonine, it is not
cited yet. That is because this AA was the last one discovered,
being described only in 1935 by the North American chemistry
William Cumming Rose.
72
Table 1 - Amino acids that have been demonstrated to be products of the

hydrolysis of proteins
Earliest observation of the amino
Earliest observation of the amino
Amino Acid Amino Acid acid as a product of hydrolysis of
acid
proteins
1 Cysteine Wollaston 1810 Glycine Braconnot 1820
2 Leucine Proust 1819 Leucine Braconnot 1820
3 Glycine Braconnot 1820 Tyrosine Bopp 1849
4 Aspartic acid Plisson 1827 Serine Cramer 1865
5 Tyrosine Liebig 1846 Glutamic acid Ritthausen 1866
6 Alanine Strecker (synthesis) 1850 Aspartic acid Ritthausen 1868
7 Valine von Gorup-Besanez 1856 Phenylalanine Schulze and Barbieri 1881
Weyl 1888
8 Serine Cramer 1865 Alanine
(Schützenberger 1879?)
9 Glutamic acid Rittausen 1866 Lysine Drechsel 1889
10 Phenylalanine Schulze 1879 Arginine Hedin 1895
11 Arginine Schulze 1886 Iodogorgoic acid Drechsel 1896
Kossel
12 Lysine Drechsel 1889 Histidine 1896
Hedin
13 Iodogorgoic acid Drechsel 1896 Cysteine Mörner 1899
Kossel
14 Histidine 1896 Valine Fischer 1901
Hedin
15 Proline Willstätter (synthesis) 1900 Proline Fischer 1901
16 Tryptophane Hopkins and Cole 1901 Tryptophane Hopkins and Cole 1901
17 Oxyproline Fischer 1902 Oxyproline Fischer 1902
18 Isoleucine Ehrlich 1903 Isoleucine Ehrlich 1903
19 Thyroxine Kendall 1815 Thyroxine Kendall 1915
20 Oxyglutamic acid Dakin 1918 Oxyglutamic acid Dakin 1918
21 Methionine Mueller 1922 Methionine Mueller 1922
Adapted from Vickery & Schmidt (1931).
THE INDUSTRIAL AMINO ACIDS
As is well known, in 1908, Dr. Kikunae Ikeda, professor of

Tokyo Imperial University, found that monosodium glutamate
(MSG) was the umami substance contained in a type of kelp
(konbu in Japanese) that had traditionally been used as soup
stock in Japanese cooking (washoku) (Ikeda, 1908). He then
succeeded in producing MSG from acid hydrolysates of wheat
or soybean proteins. Saburosuke Suzuki, the progenitor of
Ajinomoto Co. Inc., further developed this production process
and commercialized MSG as a seasoning under the brand
name of “AJI-NO-MOTO®.” The process was the first
industrialization of amino acid production and continued
successfully for approximately 50 years. Later, the production of
MSG trough the fermentative process was introduced, and since
then this production methodology has been carried out
successfully.
Although the production of amino acids dated from late
1900s, the application of industrial amino acids for feed has only
roughly 60-year history. In the late 1950s, DL-Methionine,
produced by chemical synthesis, began finding its way into
poultry feed. Production of L-Lysine by fermentation was started
in Japan during the 1960s. In addition to these amino acids, L-
Threonine and L-Tryptophan were introduced in the late 1980s.
About 20 years after was launched the L-Valine. With progress
in biotechnology, the cost of production of each amino acid has
been significantly reduced, which has been one of the key
factors in the expansion of use of amino acids in feeds. Amino
acids for feed now play very important roles in improving the
efficiency of protein utilization in animal feeding.
Kidd et al (2013) published a review about the history of
research reports and computer least cost formulation advances
that have led to present day amino acid (AA) use in poultry
diets. The authors highlighted by decade as follows:
 1950s: The introduction of least-cost formulation.
 1960s: Industry acceptance and use of least-cost

formulation.
 1970s and 80s: Mainframe systems to personal DOS
systems.
 1990s: L-Threonine and networks to Windows.
 2000s: The BCAA and the death of DOS.
It’s interesting to note that the evolution in AA usage in

feeds is linked to advances in many areas as biotechnology (for
industrial AA production), researches on AA requirements by
Universities, and the technology itself (computer advances) that
decreased runtime of linear algorithms, coupled with linear
programming formulation software advances.
These factors allowed the feed formulation with inclusion of
each subsequent AA, which provides a diet closer to the bird’s
requirement, thereby reducing AA overages and leading to a
reduction in bird nitrogen excretion. However, as each industrial
AA has reached the marketplace, there has been a period of
uncertainty toward it full adoption within practical formulation (Kidd
et al, 2013). A good example of that is the L-Valine, a feed grade
AA commercially launched in 2009 and that is still rapidly growing
it’s use in broilers diets as L-Lysine and L-Threonine are
nowadays.
Historically it was only the economic incentive which
resulted in the use of industrial amino acids in feed formulation.
However, there has been a gradual evolution and more emphasis
is now being given to supplement amino acids in terms of
sustainability and total nutrient supply. In practice, diets continued
to be dominated by the concept that formulations were considered
in terms of ingredients, but new approaches are focusing on
dietary nutrient supplying independently of the ingredients per se.
AMINO ACIDS REQUIREMENTS FOR BROILERS
It is well know that AAs requirements can vary accordingly to

many factors, as animal’s age and sex, environmental conditions
(temperature, flock density etc.), sanitary status and even when
analyzed by different mathematical methodologies.
Apart these factors, we still have to consider that the animals
are in constant changes by the genetics companies. The genetic
houses are able to change an entire broiler strain in only about four
to five years, meaning that researches on AAs requirements for that
specific strain should be done all over again from the first limiting
amino acids. According to Bourne (2007) cited by Brandalize (2016),
improvements on live weight (+50g), feed conversion ratio (-0.02
point), carcass yield (+0.1%) and breast meat yield (+0.3%) are
expected annually only considering the genetic changes. This
evolution is very clear when the look at the works of Havenstein et al.
(2003a,b) and Zuidhof et al. (2014).
Havenstein et al. (2003a,b) summarized the data from two
broiler studies carried out in 1991 and 2001. The data show that the
modern broiler in the year 2001 was nearly five times as large at 42
and 56 days of age as the 1957 randombred broiler (ACRBC), and
that the increase in body weight over the 10-year period from 1991 to
2001 was 49.9 and 81.6 grams/year at those two ages, respectively.
Edible carcass yield (Figure 1) has increased by 12.3 and 13.6 % at
42 and 56 days of age in the 2001 birds in comparison with the yield
of the 1957 ACRBC. The data from this study corroborate other
studies from Sherwood (1977) and Havenstein et al. (1994) that
consistently show that about 85-90% of the change in growth rate
has been due to genetic selection, and only 10-15% of the change is
due to improvements in nutrition and nutritional management.
Because of these changes in growth rate, the feed conversion
of broilers at a given age has dropped dramatically over the past 45
years. The trial data projected that the modern broiler in 2001
reached 1,800g body weight (BW) at about 32 days of age with a
feed conversion ratio of 1.46, while the ACRBC would have needed
an additional 17 days to reach the same BW, and its feed conversion
at that age would have been approximately 4.42. Thus, genetics,
nutrition and other management changes over the 44 year period
from 1957 to 2001 resulted in a broiler that requires approximately
1/3 the time and 1/3 the amount of feed to produce a 1,800g broiler.
Figure 1 - Broiler carcasses from the Ross 308 and the Control
(ACRBC) broilers in the 2001 study (Havenstein et al.,
2003a,b).
Zuidhof et al. (2014) found similar results when compared

two unselected meat control strains representative of broilers in
1957 (AMC-1957) or 1978 (AMC-1978) with a commercial broiler
of 2005 (Ross 308) to examine changes in growth, efficiency, and
yield resulting from commercial selection pressures. The figures
2A,B shows the results in terms of body weight, while the Figure 3
compares the age-related size of each strain.
Figure 2 - Absolute (panel A) and relative (panel B) body weight of

mixed sex University of Alberta Meat Control
unselected since 1957 and 1978, and Ross 308 broilers
(2005).
Figure 3 - Age-related changes in size of University of Alberta

Meat Control strains unselected since 1957 and 1978,
and Ross 308 broilers (2005). Within each strain,
images are of the same bird at 0, 28, and 56 d of age.
a. Lysine (Lys)
The lysine is an essential AA for the broiler maintenance,
growth and production, having as main function the synthesis of
muscular protein.
To know the lysine requirement is a base to have an
adequate and well balanced diet in terms of amino acids, because
the lysine is used as reference in the ideal protein concept. In that
sense, the incorrect consideration of lysine requirement will affect
the quantities of other AAs in the diet and that may jeopardize the
broiler’s performance.
The modern broiler strains need a higher AA content, as
can be seen in the Graphic 1. This graphic shows the broilers
requirements for digestible lysine published by Rostagno et al.
(1983, 1996, 2000, 2005 and 2011). It can be noted a great
increment in the requirements from the first to the most recent
publication, corresponding to 33, 18, 12 and 16% for the pre-
starter (1-7d), starter (8-21d), grower (22-35d) and finisher (36-
42d) phases, respectively.
Graphic 1 - Evolution of digestible lysine1,2 recommended for

broilers by Rostagno et al 4.
1
Recommendations were corrected to same levels of metabolizable energy in each
phase, being considered: ME pre-starter = 2,950 kcal/kg, ME starter = 3,000 kcal/kg,
ME grower = 3,100 kcal/kg and ME finisher = 3,150 kcal/kg.
2
The recommendations of year 1983 were calculated considering that digestible lysine
= 90.5% of total lysine.
3
Rostagno et al. (1983); Rostagno et al. (1996); Rostagno et al. (2000); Rostagno et
al. (2005); Rostagno et al. (2011).
Because of the constant evolution of the animals, Almeida

(2010) made a trial to evaluate the effect of digestible lysine levels
on broilers performance. The control group received a diet with
lysine levels of 1.20, 1.05, 1.00 and 0.90% (for pre-starter, starter,
grower and finisher phases, respectively). Treatments 2 to 5
received increments of 0.05 point in lysine level, finalizing the last
treatment with digestible lysine levels of 1.40, 1.25, 1.20 and
1.10%. It’s important to cite that the other amino acids were kept
at a constant ratio to the lysine, so their levels in the diet increased
accordingly to the lysine. The researcher concluded that higher
lysine levels (treatments 4 and 5) positively affected the results of

weight gain, feed conversion and yields of breast meat and thigh.
The addition of L-lysine (and other industrial amino acids) to
the feeds helps to reduce the use of protein sources while
favorable the entry of energetic feedstuff, sparing the use of
vegetable oils or animal fat and lowering the feed cost.
b. Threonine (Thr)
Threonine is an essential AA and is found in high

concentrations in the heart, muscles, gastrointestinal tract and
central nervous system. It is required for protein formation and
maintenance of the body's protein turnover, besides helping in the
formation of collagen and elastin (Sá et al., 2007). Low threonine
content is found in grains; therefore, in grain-based diets the
supplementation of industrial L-Threonine is almost mandatory.
For broilers, threonine is the third limiting amino acid, which
reaffirms its importance in animal metabolism and its practical
application in feed formulations. This AA is an important
component of body protein and acts as precursor of glycine and
serine. Threonine becomes more important at older animals, as
the proportion of the requirement of threonine for maintenance is
high.
In addition to acting on other vital functions of the organism,
threonine is involved in physiological functions such as digestion
and immunity (Bisinoto et al., 2007). The mucus, a secretion
produced by the gastrointestinal tract, is composed mainly of
water (95%) and mucins (5%), which are glycoproteins of high
molecular weight and especially rich in threonine. It is estimated
that more than half of the threonine consumed is used at the
intestinal level for maintenance functions, being primarily used in
the synthesis of mucin (Stoll et al., 1998; Wu, 1998). The type and
amount of mucin produced in the gastrointestinal tract influence: a)
the microbiota (as a substrate for bacterial fermentation and
fixation); b) nutrient availability (via endogenous loss of mucin as
well as nutrient absorption); and c) immune function (via control of

the microbial population and availability of nutrients) (Corzo et al.,
2007a).
Corzo et al. (2007a) verified that the requirement of
digestible threonine in the diet ranged from 0.63 to 0.66% for
broiler chickens from 21 to 42 days reared in a clean environment
(floor with new wood-shaves), resulting in dThr:dLys from 61 to
64% according to different parameters. When the broilers were
reared in a dirty environment (used wood-shaves), the
requirement ranged from 0.65 to 0.70%, resulting in dThr:dLys
ratios from 63 to 68%, thus evidencing the greater importance of
threonine in situations of greater challenge to the immune system
(Graphic 2). The authors concluded that the higher Thr
requirements for broiler chickens raised in a dirty environment may
reflect the level of exposure to the different microbial load these
birds had and the consequent change in mucin production.
Graphic 2 - Requirement of dThr (as ratio of dLys) of broilers from

22 to 42d of age reared in different environmental
conditions according to different parameters
Adapted from Corzo et al. (2007a)

Comparing the two last versions of the Brazilian Tables for

Poultry and Swine (Rostagno et al. 2005, Rostagno et al. 2011),
we can note that Thr requirement (calculated as g/phase)
increased 2.8% in pre-starter, 28.6% in starter, 14.8% in grower
and 6.0% in finisher phase. However, the ideal protein ratio of Thr
is the same in both publications (65% dThr/dLys), meaning that
the increment in Thr requirement is basically due to the increases
in Lys requirement and in the feed consumption.
However, newer researches have shown that higher ratio
dThr/dLys positively affects the broilers performance. Tillman and
Dozier (2014) published a review on AAs requirements of broilers
and based on several data already published the authors
recommended an initial dThr/dLys ratio of 70% and finalizing with
68%. An interesting point is that the author commented,
“Depending upon ingredient matrix values, local conditions, etc.
these suggested ratios might differ by as much as +/- 2 points”.
This comment shows that more attention is being given to Thr
(and other amino acids) as not only building blocks for protein, but
as also considering their other functions.
c. Branched-chain amino acids (BCAA)
The BCAA group englobes the aliphatic and highly

hydrophobic AAs Valine (Val), Isoleucine (Ile) and Leucine (Leu).
These three essential AAs share the same enzymes used in their
metabolism, being this a very important point of consideration
when defining the requirements to be used in the feed formulation.
According to Champe and Harvey (1996), these AAs are
usually found inside the proteins, being responsible for their three-
dimensional structure. Only a moderate deficiency may lead to
reduced growth rate, worsening of feed conversion, and reduction
of essential protein levels in the blood (D'Mello, 2003).
The main function of valine is the formation and deposition
of the corporal protein, being found in greater concentration in the
skeletal musculature. From a review of some publications and
personal simulations in feed formulation software, it appears that

valine is clearly fourth limiting in all vegetable-based broiler feeds
with corn and/or wheat as the primary grain source. Other possible
limiting amino acids are isoleucine, arginine, tryptophan and
glycine, but that sequence largely depends on the combination of
feed ingredients available and AAs requirements used to formulate
the diet.
Knowing the fourth-limiting amino acid is important because
it will govern the extent to which the industrial amino acids can be
incorporated into the formula. Also, the correct consideration of
this AAs and it requirement is a key to succeed when using the
ideal protein profile.
Corzo et al. (2007b) determined the requirement of
digestible valine (dVal) for broilers from 21 to 42 days of age. Data
analysis showed that weight gain and breast fillet, both in weight
and yield, were influenced by the valine levels studied (Table 2).
The authors recommended a 78% dVal:/dLys ratio, or a minimum
dietary level of 0.74% of digestible valine.
Table 2 - Performance of broilers from 21 to 42d old fed with

increasing levels of dVal
dVal. Weight gain Feed conversion Breast fillet

(%) (g) (g/g) Weight (g) Yield (%)
0.59 1,223.0 2.23 318 23.9
0.64 1,291.0 2.22 341 24.5
0.69 1,352.0 2.19 350 24.8
0.74 1,382.0 2.06 357 25.2
0.79 1,390.0 2.06 360 24.8
0.84 1,385.0 2.09 358 24.8
Quadratic
0.04 0.41 0.04 0.01
reg.
dVal/dLys 78 - 77 74
Berres et al. (2010) verified that dVal/dLys of 75% to 76%

resulted in greater weight gain and better feed conversion in
growing broiler (22 to 42d). However, increasing the ratio to 78%
reduced the percentage of abdominal fat. Campos et al. (2010)
recommended the ratio dVal/dLys of 79% after observe improved
performance of growing broiler fed diets with that ratio.
Rostagno et al. (2011) recommend levels of 1.02; 0.937;
0.882 and 0.827% of digestible valine for broilers in each phase
(pre-starter, starter, grower and finisher), respectively, with
dVal/dLys of 77% from 1 to 21d and 78% from 22 to 42d.
Tillman and Dozier (2014), based on a review of several
scientific studies published, reached the recommendations of
digestible valine for broilers of 0.98; 0.90; 0.84; 0.77 and 0.70% in
the diet for each phase. These values correspond to the dVal/dLys
ratios of 76, 76, 77 and 78% in the respective phases.
Talking about the Isoleucine, this AA has been considered
the fifth limiting amino acid in broilers diets based on corn and
soybean meal (Berres et al., 2010; Goulart et al., 2009). Trials to
determine the requirement of this AA have been made and the
results differ a little.
Tillman and Dozier (2014) made a review of several papers
and recommended the dIle/dLys ratios of 66 (until 14d old), 67
(from 15 to 28d) and 68% (until 42d old) for broilers. However, the
data analyzed by these researchers showed a wide range of
results, indicating Ile requirements (as ratio of dLys) from 62% up
to 75%. That vary can be partially explain by differences in the
strain studied, the statistical method applied for data analysis, the
parameter analyzed (performance or carcass) etc. The definition
of a requirement to be used for feed formulation should be related
to the objectives of each company, for example, a better feed
conversion or a higher breast yield.
Those numbers are very similar to the ones recommended
by Rostagno et al. (2011), which are the dIle/dVal ratios of 67%
(until 21d old) and 68% for broilers.
The other BCAA is the Leucine. Although it’s also an

essential AA, the content of Leu in common broiler diets is always
higher than the requirement, so there’s no need to concern about
supplementing L-Leu. On the other hand, this excess in the diets
should be closely monitored due to the metabolism pathway the
three BCAAs share. Since AAs cannot be store, any excess will be
catabolized. The problem identified in case of BCAAs is that the
second step of their catabolism is under the influence of Leu,
which activates the BCKDH enzyme complex at the liver leading to
the catabolism of Val and Ile when Leu is in excess. Even if the
Val and Ile are deficient in the diet, the excess of Ile will enhance
the catabolism of all three AAs. D’Mello and Lewis (1970)
observed in growing broilers that an excess of Leu lowered
circulating levels of Ile and Val, while the excess of Val had no
effect on birds’ response to Ile.
Maia et al. (2012) evaluated the effect of regular and high
dietary levels of Ile, Leu and Val on the performance of growing
broilers. The authors found that the increase in dLeu/dLys ratio
from 107% to 150% in broiler feeds reduced weight gain (WG) and
worsened feed conversion ratio. However, the increase in the
ratios of dIle/dLys (from 67% to 80%) and dVal/dLys (from 77% to
90%) did not influence broiler performance. Another information
from this work that is interesting to highlight is the positive effect of
higher dVal on the birds’ weight. With all three BCAAs within
recommendation (Rostagno et al., 2011), the broilers had a WG of
630.4g. Once the Leu was in excess, the WG dropped to 608.9g
(reduction of 3.4%). Now with the higher dVal/dLys ratio, the
excess of Leu depressed the WG only by 0.17% (from 627.6g vs
626.5g), showing that this higher supply of dVal was able to
assure some free valine to be deposit as protein (meat).
d. Other amino acids
Some of the called nonessential AAs (NEAA) for broilers

play important roles in the body. For example, glutamine (Gln),
glutamate (Glu), and arginine (Arg) acts regulating gene

expression, cell signalling, antioxidative responses, and immunity.
Additionally, glutamate, glutamine and aspartate (Asp) are major
metabolic fuels for the small intestine and they, along with glycine
(Gly), regulate neurological function. In the sequence, we are
presenting some trials made with these amino acids.
Murakami et al. (2012) studied the effects of a starter diet (1
to 21d) supplemented with L-Arg on the production performance
and intestinal mucosa morphometry of broilers. They
supplemented five levels of L-Arg to achieve the ratios dArg/dLys
from 110% to 142%. It was observed that L-Arg supplementation
increased the live weight while feed intake of the birds didn’t
change, and so lowering the feed conversion ratio. Also the
duodenum villus:crypt ratio increased and the crypt depth
decreased in the first week in response to increasing dietary Arg.
The authors concluded that broiler Arg dietary supplementation in
the starter diet improved production performance and small
intestine morphometry, especially in the first week.
Olubodun et al. (2013) evaluated the effect of dietary
supplementation of L-Gln and L-Glu acid on acute phase protein
and heat shock protein 70 of broilers in heat stress. Based on the
results of HSP70 expression, the authors concluded that dietary L-
Gln and L-Glu supplementation could be beneficial in enhancing
heat tolerance in broiler chickens.
Olubodun et al. (2015) designed another study to
investigate the effect of L-Gln and L-Glu supplementation on the
performance, intestinal morphology and amylase activity in broiler
chickens under the hot and humid tropical conditions. The results
from this study showed that supplementing diets with L-Gln and L-
Glu may improve growth performance, survivability, intestinal
morphology and amylase activity of broiler chickens under heat
stress condition.
Manvailer et al. (2015) evaluated plans of L-Gln + L-Glu
supplementation to broilers from one to 42 days old on hot
environment. Six hundred and ninety male birds Cobb were
distributed in a randomized block design with five plans of

supplementation (considering phases and supplementing
quantity). They concluded that for birds under high temperatures, it
is recommended to supplement 1.0% of L-Gln + L-Glu from 1 to
14 days of age and then 0.5% until 21 days, what resulted in
higher weight gain, carcass weight and lower feed conversion.
As more supplemental AA enter formulation, a concomitant
decrease in soybean meal occurs. In this scenario, and in
particular all vegetable diets, glycine (Gly) seems to be
semiessential deeming a Gly + Serine (Ser) minimum to be set in
least cost formulation (Kidd et al, 2013).
The problem is that there is no consensus about the
requirement of Gly or Gly+Ser. For example, Rostagno et al.
(2011) informed the ratio of dGly+dSer/dLys of 147% (until 21
days old) and 137% for later phases as requirement. However, Wu
(2014) suggests the ratio of 245% as optimal for broilers until 56
days old. Besides this difference in recommendation, what is
known is that Gly has numerous important function, as a precursor
of glutathionine peroxidase, nucleic acids, creatine, heme, bile,
and it serves as a methyl source [58]. Moreover, each molecule of
uric acid contains one molecule of Gly, and under some
circumstances, Gly is considered essential.
Dean et al. (2006) conducted six experiments to determine
the effects of low crude protein (CP) in diets for broilers and to
evaluate limiting essential and nonessential amino acids in these
diets. In the sixty trial, chicks were fed the positive control (CP of
22.2%) or the low CP diet (16.2%) containing from 1.80 to 3.00%
Gly + Ser with increments of 0.15 points. Glycine addition to the
low CP diet increased G:F linearly (P <0.001). In summary, the
authors concluded that low CP diets result in optimal growth of
broilers with Gly + Ser levels of 2.44%.
Yuan et al. (2012) designed a study to evaluate the
possibility of using additional supplemental amino acids in diets for
young broiler chicks and to evaluate the effect of adding Gly to
low-CP diets. The first diet was formulated allowing only a Met
supplement; as each succeeding amino acid became limiting, that

amino acid was also provided in supplemental form. In order, the
limiting amino acids were Met + Cys, Lys, Thr, Val, Ile, Arg, Trp,
and His. The diets from Thr addition until Hys addition were
divided in two aliquots and Gly was added to one of them to
provide a minimum of 2.0% Gly + Ser. This resulted in 14
experimental diets. This work showed that diets low in CP resulted
in decreased BW of 18d old broilers when the Gly + Ser level fell
below 1.71% and increased the feed conversion at levels lower
than 1.87%. However, the addition of Gly to these diets to a
minimum level of 2.0% significantly improved performance to
equal that of diets fortified with the amino acids Met, Lys, and Thr.
Therefore, Gly may be critical in the use of low-protein amino acid
supplemented diets for broilers.
Efforts are made continuously by universities, research
institutions and industry in order to update the requirements of
AAs for broilers and their ratios to digestible lysine. Table 2
presents different profiles of the ideal protein for broiler chickens
according to the recommendations of Rostagno et al. (2011), Wu
(2014), Ajinomoto Eurolysine (2015) and Ajinomoto do Brasil.
Table 3 - Ideal protein profile recommended for broilers
Rostagno et Ajinomoto Ajinomoto do

Wu (2014)
al, (2011) Eurolysine Brasil
Phase (days) Phase (days) Phase (days) Phase (days)

Digestible
amino acid 1-21 22-56 1-21 22-56 1-21 22-56 1-21 22-56
Lysine 100 100 100 100 100 100 100 100
Met + Cys 72 73 72 75 75 75 72 73
Threonine 65 65 67 70 67 67 65 67
Valine 77 78 77 80 80 80 78 78
Isoleucine 67 68 67 69 67 67 67 68
Arginine 108 108 105 108 105 105 108 108
Tryptophan 17 18 16 17 17 17 17 18
Hystidine 37 37 35 35 36 36 36 36
Phe + Tyr 115 115 105 105 105 105 105 105
Leucine 107 108 109 109 105 105 107 108
2. The future of AA nutrition
With the advances in scientific research in the area of

animal nutrition and metabolism, as well as in the production
technology of AAs, it is becoming possible to formulate feeds with
lower protein content and levels of AAs closer to the needs of the
animal. The closer the AA composition of the diet is to the animal
requirement, the more efficient will be the use of the supplied
protein, and that positively reflects on the use of the other
nutrients.
Table 4 shows examples of formulations made using the
concept of the ideal protein profile. In this example, it was
considered that the industrial amino acids L-Tryptophan, L-
Isoleucine and L-Arginine are also available at accessible prices to
be used in broilers diets, so the least-cost formulation software
automatically used them. In the first column a common diet used
nowadays with the inclusion of only DL-Met, L-Lys and L-Thr,
although L-Val is already available at the market since the year
2009. Moving to the columns at right, we can see the effect of
including up to seven industrial AAs on the nutritional and
centesimal composition of this growing broiler diet. It is possible to

note that:
 After the well determined first three limiting AAs (Met, Lys
and Thr), the inclusion of new industrial AAs followed the
sequence: L-Valine, L-Isoleucine, L-Tryptophan and L-
Arginine, what represents the limiting order (from the 4 th to
7th, respectively) in this type of diet;
 Considering this example, the 8th limiting amino acid were
Phenylalanine + Tyrosine;
 Related to the nutrients, it’s good to emphasize the reduction
of CP content in the diets, which reduced from 21.0% to
18.6%;
 At the bottom of the formulation are the AAs ratios to lysine,
where can be noted that the addition of industrial AAs
provides a better adjustment of the diet nutrients with relation
to the requirements of AAs in the ideal protein profile
(increment in the green area).
An important point to be highlight is the nitrogen balance,

cited in the last line of table 4. As the CP of the diets reduces,
there is a possibility of lack of non-essential nitrogen (NEN) and so
this may became limiting. D’Mello (2003) said that a definition of
an optimal ratio between essential nitrogen (EN) and total
nitrogen (TN) in low protein diets could be very important to
achieve better performance and efficiency of protein utilization.
Published works with rats (Heger, 1990; Stucki e Harper,
1962), broilers (Bedford e Summers, 1985; Stucki e Harper,
1961), turkey (Bedford e Summers, 1988) and swine (Wang e
Fuller, 1989) suggested that the optimal growth rate happened
when the ratio EN:TN was in the range of 40 to 65%.
Table 4 - Example of feed formulation considering the inclusion of

new industrial amino acids
Broilers – grower 1 phase

DL-Met + L- DL-Met + L-Lys
DL-Met + DL-Met + DL-Met +
Lys + L-Thr + + L-Thr + L-Val
Ingredients L-Lys + L-Lys + L- L-Lys + L-Thr
L-Val + L-Ile + + L-Ile + L-Trp +
L-Thr Thr + L-Val + L-Val + L-Ile
L-Trp L-Arg
DL-Methionine 0.265 0.282 0.296 0.308 0.336
L-Lysine HCl 0.122 0.185 0.236 0.282 0.383
L-Threonine 0.030 0.058 0.080 0.101 0.145
L-Valine - 0.034 0.061 0.085 0.139
L-Isoleucine - - 0.028 0.054 0.112
L-Tryptophan - - - 0.008 0.025
L-Arginine - - - - 0.118
Corn 56.754 59.157 61.052 62.797 66.456
Soybean meal 45% 32.341 30.222 28.536 26.981 23.600
Soybean oil 4.316 3.859 3.483 3.133 2.383
Meat & bone meal 41% 4.619 4.646 4.669 4.689 4.736
NaCl (salt) 0.425 0.424 0.424 0.424 0.423
Limestone 37% 0.728 0.732 0.735 0.738 0.744
Premix 0.400 0.400 0.400 0.400 0.400
Total 100.000 100.000 100.000 100.000 100.000
Nutritional Values
Crude Protein (%) 21.00 20.34 19.84 19.37 18.58
Met. Energy (kcal/kg) 3150.0 3150.0 3150.0 3150.0 3150.0
Dig. Lysine (%) 1.10 1.10 1.10 1.10 1.10
Calcium (%) 0.84 0.84 0.84 0.84 0.84
Av. Phosphorus (%) 0.42 0.42 0.42 0.42 0.42
Dig. AAs / Dig. Lys ratios
Met+Cys / Lys 0.73 0.73 0.73 0.73 0.73
Threonine / Lys 0.65 0.65 0.65 0.65 0.65
Valine / Lys 0.78 0.78 0.78 0.78 0.78
Isoleucine / Lys 0.71 0.68 0.68 0.68 0.68
Tryptophan / Lys 0.20 0.19 0.18 0.18 0.18
Arginine / Lys 1.22 1.16 1.12 1.08 1.08
Phe+Tyr / Lys 1.39 1.33 1.28 1.24 1.15
Hystidine / Lys 0.45 0.43 0.42 0.40 0.38
Leucine / Lys 1.45 1.41 1.37 1.34 1.27
Nitrogen balance
EN:TN - 46.5% 47.8% 48.9% 51.2%
AAs in Ideal
excess Protein
In another work, Heger et al. (1998) evaluated the effect

of EN:TN ratio on the protein utilization by growing pigs. They
tested 6 ratios (from 25 to 86%) being the diets formulated with
the same EN but varying the TN. The authors concluded that
optimization of the EN:TN value may substantially contribute to

the reduction of N excretion. They commented that in a diet
supplying sufficient quantities of essential amino acids allowing
rapid growth, the N excretion may be reduced by decreasing
the concentration of total N up to the point at which
nonessential N becomes limiting. The breakpoint in N retention
was identified in this experiment at an EN:TN value of 0.48
(Figure 4).
Figure 4 - The relationship between essential nitrogen:total

nitrogen (EN:TN) value and nitrogen retention at
constant concentration of total nitrogen in the diet.
(Adapted from Heger et al., 1998).
More recently, Maia et al. (2015) evaluate the effect of

two EN:TN ratios (51.5 and 47.7%) on performance of growing
broilers (8 to 21 days old). The authors concluded that broilers
fed diet with the lower ratio studied had better weight gain and
feed conversion rate than the broilers of other treatment. In a
sequence of this work (data not published), the authors decide
to evaluate five ratios for broilers at the same age of from first
work. With this second work they concluded that the best
performance was observed in broilers fed diets with EN:TN
ratio up to 50%.
These findings show us how important is to have non-
essential amino acids in the diets (as source of NEN) in order
to achieve an optimal growth and protein deposition.
Making a link of these more recent works with broilers
and the diets formulated in table 2, we can see that EN:TN ratio
should not be a concerning point in the actual commercial diets
utilized. Moreover, we can consider that is still space to add
two more amino acids after L-valine (or that we can work with
up to six industrial AAs in the diets) and even thou the EN:TN
will be in the range so far considered optimal.
CONCLUSION
The challenge to feed the population in the near future is

big and the broiler industry plays an important role on this.
Without the industrial amino acids it would be very difficult, if
not impossible, to produce the quantity of meat, milk, fish and
eggs demanded by world consumers. The availability of amino
acids has allowed to produce feeds using smaller quantities of
protein rich raw materials allowing these limited scarce
resources to be used more sparingly.
The supplementation of industrial amino acids is
essential for the formulation of feeds based on the ideal protein
concept, and as more industrial amino acids become available,
the easier and more accurate will be the fulfillment of the
established nutritional requirements.
Together, the development of new technologies and the

continuous studies on broiler nutrition and production are
utmost important to guarantee the supply of animal protein.
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NUTRITIONAL RECOMMENDATIONS FOR LAYING
HENS AND JAPANESE QUAILS
§, ¥,
Fernando Guilherme Perazzo Costa ⃰, Matheus Ramalho de Lima ⃰, Danilo
§, §, €,
Cavalcante ⃰, Guilherme Souza Lima ⃰, Danilo Vargas Gonçalves Vieira ⃰
§
Federal University of Paraiba, Areia, Paraíba, Brazil
¥
Federal University of Southern Bahia, Teixeira de Freitas, Bahia, Brazil
€
Federal University of Tocantins, Araguaína, Tocantins, Brazil
⃰Study Group on Poultry Technologies
ABSTRACT
The nutritional requirements of laying hens and Japanese

quails are constantly updated and each year new studies are
published, leading to increased dietary efficiency and poultry
productivity. Therefore, it is necessary to update these nutritional
recommendations observing the main changes of these
requirements.
INTRODUCTION
Constant advances have been observed in poultry

production, particularly for laying hens. Increase in the number of
eggs per hen housed and the reduction in age at the first egg are
the main characteristics of this progress. These results come from
interrelated factors such as genetics, nutrition, health and
management that, if well done, maximize poultry performance and
increase farm profitability.
At slower steps, quail farming has growing recent years.
The diets for Japanese quails were formulated based on the
recommendations for laying hens. However, research has now
been conducted to determine the requirements of these birds at
different stages, which has resulted in greater farmers’
satisfaction, allowing bigger investment in this sector.
Studies have shown that changes in nutritional
requirements caused by genetic selection, both for laying hens
and Japanese quails, have influenced amino acid, energy and

mineral requirements, resulting in new nutritional
recommendations for these birds. For these reasons, it is
necessary to survey and update the nutritional recommendations
for laying hens and Japanese quails published in recent years.
NUTRITIONAL RECOMMENDATIONS FOR LAYING HENS
Changes in nutritional requirements caused by genetic

selection and breeding mainly influenced amino acid and energy
requirements, resulting in new nutritional recommendations for
laying hens, along with modifications in patterns and genetic
determinants.
The concept of ideal protein was developed to balance the
amount of amino acids provided to maximize its use in animal
metabolism without excess or lack in the body. Once this concept
was established, feed costs were reduced by decreasing crude
protein (CP) levels. It started the industrial production of amino
acids. Along with this evolution, researchers have studied
specifically each amino acid, how they act on metabolism and how
to use them in feed. This determined the order of amino acid
limitations as a function of the source of the ingredients used in
the diets.
Methionine (Met) is considered the first limiting amino acid
for laying hens in diets composed of soybean meal and corn and it
is usually supplemented using DL-methionine from industrial
origin. In metabolism, Met participates in protein synthesis. It is the
precursor of cysteine and a methyl donor. It also plays a role in the
production of immunoglobulin G, directly contributing to increase
the resistance of birds to diseases. As Met is an active amino acid
in the immune system, its requirement can be modified according
to the sanitary challenge of the poultry farm.
D'Agostini et al. (2012a,b) recommended levels of
methionine+cystine (Met+Cys) for replacement hens, light and
semi-heavy, from 7 to 12 weeks old, of 0.710% Met+Cys (0.639 %
digestible Met+Cys) and 0.706% Met+Cys (0.635% digestible
Met+Cys), respectively. For the 13th to 18thweek, the

recommendation is0.679% total Met+Cys (corresponding to
0.611% digestible Met+Cys) for lightweight birds and 0.646% total
Met+Cys (corresponding to 0.581% digestible Met+Cys) for semi-
heavy hens.
Lysine (Lys) is the second limiting amino acid. As such, it is
used as the reference amino acid, i.e., Lys is taken as 100% and
all other amino acids are supplemented in proportion to it. It
actively participates in the muscular synthesis and the requirement
of this amino acid is higher in the first stages of life of laying hens.
It is well known amino acid and its concentration in the diet for
laying hens can be predicted using mathematical equations that
take into account the average daily intake of the bird, the weight,
the daily weight gain and egg mass.
Araujo et al. (2014) recommended intake of 202, 300 and
146, 312 and 259 mg/day of Lys in stages from 2 to 6, 8 to 12 and
14 to 18 weeks of age, respectively, for better weight gain and
feed conversion of DeKalb lightweight birds. The W-36 line
management guide (Hy-Line, 2015) recommends a digestible Lys
level of 1.05, 0.98, 0.88, 0.76 and 0.78% from 1 to 3, 3 to 6, 6 to
12, 12 to 15 and 15 to 17 weeks of age.
Threonine (Thr) is another essential amino acid for laying
hens that is found in high concentrations in the heart, muscles,
skeleton and central nervous system. It actively participates in
protein synthesis and maintenance of protein turnover and
collagen synthesis (Sá et al 2007). Thr is involved in other
physiological functions, including digestion and immunity (Bisinoto
et al., 2007). The mucous secretion produced by the
gastrointestinal tract is mainly composed of water (95%) and
mucins (5%), which are high molecular weight glycoproteins
especially rich in threonine.
In relation to the recommendations of digestible Thr, Lima
et al. (2015) recommended0.597% for first cycle laying hens, with
Thr:Lys ratios varying from 56 to 81. Meanwhile, Cardoso et al.
(2014) recommended a Thr:Lys ratio of 75 for lightweight birds at
the second laying cycle. The Hy-Line management guide (2015)

recommends 0.55% and 0.51%, respectively, for these birds.
Tryptophan (Trp) plays important roles in the metabolism of
birds as a precursor to niacin and serotonin. Due to its importance
in the immune system and its metabolites are the main products
related to the regulation of intake and stress, it is essential to
establish the correct level of this amino acid in the feed. Trp
deficiency causes a reduction in feed intake (Peganova & Eder,
2002) and its requirement can be influenced by long chain neutral
amino acids present in the diets, such as isoleucine, valine,
leucine, phenylalanine, tyrosine, Met and histidine. The amino acid
L-Tryptophan from industrial origin is an alternative form of
supplementation.
Valine (Val) may be considered as one of the potential
limiting amino acids for birds. This limitation is particularly evident
for older birds when dietary protein decreases and energy content
increases. The Brazilian Tables of 2011 recommend for
lightweight birds (around 1,600 kg)654 mg/day of digestible Val
and 735 mg/day of total Val. For semi-heavy laying hens (around
1,800 kg), 675mg/day of digestible Val and 759 mg of total Val/day
are recommended. Lelis et al. (2014), in more recent studies with
semi-heavy laying hens, recommend 567 mg of digestible Val/day.
Arginine (Arg) is an amino acid that has aroused interest
due to its antagonistic relation to Lys. Excess dietary Lys reduces
Arg levels in the body because these amino acids compete for the
same site of absorption; in addition, excess Lys stimulates renal
arginase, further reducing Arg levels. Carvalho et al. (2015)
reported that increasing dietary levels of digestible Lys and Arg
reduced eggshell quality and albumin solids, respectively. The
recommended amino acid level to improve egg quality is 700 mg
for both digestible Lys and digestible Arg/kg of feed,
corresponding to a digestible Lys:digestible Arg ratio of 100.
Isoleucine (Ile) is an essential amino acid needed for the
normal growth and development of laying hens, as well as egg
production and egg mass. It is a branched chain amino acid such
as Val and leucine. Studies conducted by Rocha et al. (2013)

demonstrate that the best digestible Ile:digestible Lys ratio for Hy-
Line W36 laying hens at 24-40 weeks of age is 0.84:1,
corresponding to 681 mg Ile and 811 mg Lys/day/hen.
In addition to amino acids, there are studies to determine
the nutritional recommendations for laying hens in relation to
metabolizable energy levels. The ideal level of energy that each
feed should contain is established using mathematical equations,
taking into account the body weight of the birds, weight gain in
grams per day, egg mass and average temperature in °C. Another
factor that determines the requirement of birds is the ideal
relationship between the amount of energy in the diet and the
levels of all other nutrients. This relationship is extremely important
to ensure satisfactory nutrient intake, leading to efficient egg
production.
The nutritional requirements of laying hens are also
influenced by their feed intake capacity. Several situations
contribute to deprived feed intake, which reduces nutrient supply
to the body, leading to decreased performance. A classic situation
of low feed intake is found in hot climate regions, since high
temperatures inhibit feed intake. Consequently, adjustments in diet
should be made. The nutritionist must adjust the dietary energy
level, reducing heat increment and ensuring that the levels of other
nutrients meet the requirements of the animal in the necessary
quantity, even with low feed intake. Such procedures are only
possible with the introduction of new ingredients in the feed, such
as oils and amino acids from industry.
Studies have been developed to supply the right amount of
nutrients the animal needs to produce efficiently. These
adjustments in the diet, besides contributing to the costs of
production and poultry productivity, are directly related to the
preservation of environment and the sustainability in the poultry
activity. In this sense, the requirements of minerals for laying hens
have been the focus of many studies.
Calcium (Ca) is the most abundant mineral in the birds’

body. It is considered one of the main bone constituents and it
plays a fundamental role in the control of the cellular functions of
nervous and muscular tissues, as well as the hormonal activities of
blood coagulation, skeletal formation and eggshell structure.
Phosphorus (P), on the other hand, is the mineral that
accompanies the metabolism of Ca, mainly in terms of absorption
and serum levels. It is involved in the collagen synthesis and bone
mineralization, increasing the resistance of bones and accelerating
fracture healing - activating coenzymes for the vitamin B-complex
functioning -, in addition to participate as a buffer in intracellular
medium and tubular fluids of the kidneys.
Ca and P are indispensable for egg production and quality,
since calcium is the main component of the eggshell and
phosphorus is fundamental for its satisfactory mineralization and
calcification. The efficiency of use of these minerals is closely
related to the quantity and relationship between them. Excess Ca
in the bloodstream is harmful and decreases the absorption of
other minerals. The same happens with P, because its excess
negatively affects calcium mobilization from bone and the eggshell
mineralization.
CHANGES INTHE RECOMMENDATIONS OVER TIME
The nutritional recommendations for laying hens are

constantly updated, especially because their productive capacity
changes over time. The importance of proper nutrition for poultry is
critical for an efficient egg production, especially focused
nutritional efficiency, to convert feed into an extremely rich and
functional food, the egg.
Regarding the nutritional recommendations for lightweight
and semi-heavy laying hens, it is fundamental to observe and
adapt the diets prior to the productive phase. Much is known about
the importance of this period and the impact in the productive life
of the laying hen when the specific nutrition in this phase is fully
attended. In this context, we systematized data from the

management guide for commercial lineages of lightweight and
semi-heavy hens, in order to compare the changes in nutritional
recommendations.
Regarding the recommendations for the same lineage in
management guides of 2009 and 2015 (Table 1), over a relatively
large period considering poultry production cycle, there was only
changes in nutritional recommendations for crude protein (CP),
Sodium (Na) and chlorine (Cl). These minerals were reduced only
from4 to 6 and from 7 to 12 weeks. In contrast, a new lineage had
similar nutritional management, but with recommendations varying
considerable, from increases to decreases in relation to the
previous lineage with recommendations in a management guide of
2015.
The data show an increase in calcium in the pre-laying
phase by 0.2 pp (percentage points) with the changes for the new
lineage compared to that recommended for the other lineage in
2015. There were also increases in sodium and chlorine in the
phases up to 12 weeks and indigestible Met+Cys from 0 to 3 and
7 to 12 weeks of age, although the previous changes in
recommendations always decreased its dietary level (Table 1).
Table 1 - Variation in nutritional recommendations, per unit, for

commercial LIGHTWEIGHT LAYING HENS in 2009
and 2015 and for a modern lineage in 2015, from 0 to
17 weeks old.
Prelay
0-3 w 4-6 w 7-12 w 13-15 w
(16-17w)
1 White 1 White 1 White 1 White 1 White
09 2 3 09 2 3 09 2 3 09 2 3 09 2 3
% 15 16 15 16 15 16 15 16 15 16
PB 20,0 19,0 -0,75 18,0 -0,50 17,0 -1,0 17,0 -0,50
Ca 1,0 1,0 1,0 1,4 -0,4 2,5 0,20
Pd 0,5 0,49 0,47 0,45 0,02 0,48
K -
Na 0,18 0,03 0,18 -0,01 0,03 0,18 -0,01 0,01 0,18 0,18
Cl 0,18 0,03 0,18 -0,01 0,03 0,18 -0,01 0,01 0,18 0,18
Met+Cys 0,74 0,04 0,74 0,67 0,01 0,59 -0,02 0,66 -0,02
Lys 1,05 -0,03 0,98 -0,04 0,88 -0,04 0,76 -0,08 0,78 -0,04
Trp 0,18 0,18 -0,01 0,17 0,15 0,16
Thr 0,69 -0,03 0,66 -0,05 0,60 -0,04 0,52 -0,05 0,55 -0,04
Ile 0,74 -0,02 0,71 -0,04 0,65 -0,03 0,57 -0,06 0,62 -0,04
Val 0,76 -0,02 0,73 -0,04 0,69 -0,03 0,61 -0,06 0,66 -0,03
Arg 1,12 -0,06 1,05 -0,07 0,94 -0,07 0,81 -0,10 0,83 -0,06
¹Recommended level according to the management guide for commercial lineages of
lightweight birds in 2009; ²Variation in nutritional recommendation for the commercial
lineage of laying hens in 2015; ³Variation in nutritional recommendation for a new
commercial lineage of laying hens in 2016; *Gray spaces represent data without
variation, or without recommendation, in the case of K; **The units of the items are
%(Crude Protein), g/day(Calcium) and mg/day (others).
When we observe the recommendations for semi-heavy

laying hens until pre-laying phase (Table 2) considering the same
lineage in two moments of changes in nutritional
recommendations in 2009 and 2014, we notice an important
change for lightweight birds. However, the most observed are
increases in nutritional levels, especially for amino acids, but also
with some reductions in Ca, Met+Cys and Arg.

commercial SEMI-HEAVY LAYING HENS in 2009 and
2014from0 to 17 weeks old.
0-3w 4-6w 7-12w 13-15w Prelay
Brown Brown Brown Brown Brown

% 091 142 091 142 091 142 091 142 091 142
PB 20,00 18,25 17,50 16,00 16,50
Ca 1,00 1,00 1,00 1,40 -0,4 2,50
Pd 0,45 0,44 0,43 0,45 0,48
K
Na 0,18 0,17 0,17 0,18 0,18
Cl 0,18 0,17 0,17 0,18 0,18
Met+Cys 0,75 0,020 0,70 0,020 0,65 0,010 0,57 -0,01 0,63 -0,01
Lys 0,99 0,020 0,90 0,020 0,80 0,020 0,65 0,02 0,70 0,02
Trp 0,18 0,17 0,17 0,14 0,01 0,15 0,01
Thr 0,63 0,020 0,59 0,010 0,54 0,010 0,44 0,02 0,48 0,02
Ile 0,69 0,020 0,65 0,010 0,59 0,020 0,49 0,01 0,56
Val 0,71 0,020 0,67 0,010 0,62 0,020 0,52 0,02 0,60 0,01
Arg 1,06 -0,010 0,96 0,86 -0,010 0,70 0,75
¹ Recommended level for the lineage in 2009; ²Level with positive or negative variation
for the lineage between 2009 and 2014; *Gray spaces represent data without variation,
or without recommendation, in the case of K.
Considering the data in Table 3, when comparing

lightweight birds from 2009 to 2015, a significant reduction in the
live weight is verified, justifying a lower daily and cumulative feed
intake. When comparing the 2015 lineage with the new 2016
lineage, it is verified that the live weight was even smaller, but a
recovery of daily feed intake after the 12th week was observed. On
the other hand, cumulative feed intake remained low. When we
compare the semi-heavy hens, there is not an expressive change
in live weight, although with greater reduction along the weeks.
However, the greater cumulative feed intake per week is what
draws more attention.
In this sense, coupled to the information in Tables 1 and
Table 2, and considering the data in Table 3, the changes show a
greater tendency to reduce the nutritional levels for lightweight
laying hens and to increase the levels for semi-heavy hens. The
reason is clear, especially as the lightweight birds get lighter and
therefore consume less feed. The opposite proportionally occurred
for semi-heavy hens, which maintained the live weight, but
increased the cumulative feed intake capacity. Therefore,
lightweight hens require a downward adjustment in nutritional
levels and the opposite is necessary for semi-heavy hens.
Table 3 - Live weight (LW, g/hen), daily intake (FI, g/day) and
cumulative feed intake per week (CFI, g/w) of
lightweight and semi-heavy hens in the above-
mentioned recommendations
White Brown
LW, g/bird CFI, g/w LW, g/bird FI, g/w
20091 20152 20163 2009 2015 2016 2009 2014 2009 2014
65 -4 9 91 7 -14 70 -2 70 28
110 2 8 203 -7 -7 120 1 196 21
170 6 4 336 -14 14 200 -16 343 35
250 -5 5 539 -14 0 250 7 532 35
320 4 -4 805 -14 -49 335 14 742 63
410 3 -3 1092 -14 -91 450 -4 994 77
500 2 8 1393 -21 -112 540 3 1274 84
590 1 9 1715 -28 -133 640 10 1575 98
690 -10 20 2051 -42 -147 750 7 1918 98
790 -11 1 2408 -56 -154 860 3 2296 84
870 -2 -8 2779 -70 -154 960 0 2702 84
940 7 -27 3157 -84 -140 1070 -22 3136 84
1020 -4 -36 3549 -98 -119 1120 5 3591 98
1090 -5 -55 3948 -112 -84 1200 -7 4067 112
1160 -6 -74 4361 -126 -42 1260 1 4557 126
1200 -7 -63 4788 -140 14 1320 9 5082 126
1240 -8 -62 5222 -154 84 1400 -3 5621 133
¹Absolute data observed in management guides; ²Difference between 2015 and 2009, in the
units of each variable; ³Difference between 2016 and 2015, in the unit of each variable.
By applying the evaluation criteria on the nutritional

recommendations for hens until the pre-laying period, it is possible
to systematize these levels for laying hens, where we present the
data of the lightweight and semi-heavy layers, respectively, in
Tables 4 and 5.

commercial LIGHTWEIGHT LAYING HENS in 2009 and
2015 and for modern lineage in 2015 at peak production
until phase IV
Production peak Phase II Phase III Phase IV
White White White White

091 091 091 091
152 163 152 163 152 163 152 163
PB 16,0 15,5 15,25 15,0
Ca 4,0 0,15 0,25 4,2 0,10 0,30 4,35 0,05 0,35 4,5 0,10 0,15
Pd 500,0 -15,00 25,0 480,0 -10,0 -30,0 460,0 -10,0 -60,0 400,0 -10,0
K
Na 180,0 180,0 180,0 180,0
Cl 180,0 180,0 180,0 180,0
Met+Cys 676,0 29,0 630,0 49,0 596,0 50,0 570,0 43,0
Lys 805,0 15,0 750,0 40,0 710,0 50,0 695,0 35,0
Trp 169,0 3,0 158,0 8,0 149,0 11,0 146,0 7,0
Thr 564,0 10,0 525,0 28,0 497,0 35,0 487,0 24,0
-
Ile 636,0 -8,0 12,0 593,0 -8,0 31,0 561,0 -7,0 39,0 549,0 27,0
7,00
Val 725,0 -17,0 13,0 675,0 -15,0 35,0 639,0 -14,0 43,0 626,0 -14,00 30,0
Arg 861,0 -24,0 16,0 803,0 -23,0 42,0 760,0 -22,0 52,0 744,0 -21,00 36,0
¹Recommended level according to the management guide for commercial lineages of
lightweight birds in 2009;²Variation in nutritional recommendation for the commercial
lineage of lightweight laying hens in 2015; ³Variation in nutritional recommendation for a
new commercial lineage of lightweight laying hens in 2016; *Gray spaces represent data
without variation, or without recommendation, in the case of K; **The units of the items
are % (Crude Protein), g/day (Calcium) and mg/day (others).

commercial SEMI-HEAVY LAYING HENS in 2009 and
2014 at peak production until phase IV
Production peak Phase II Phase III Phase IV
Brown Brown Brown Brown

1 2 1 2 1 2 1 2
09 14 09 14 09 14 09 14
PB 17,0 16,75 16,0 15,5
Ca 4,0 0,20 4,40 -0,10 4,70 -0,20 4,9 -0,100
Pd 440,0 20,0 400,0 20,0 360,0 20,0 350,0 10,0
K
Na 180,0 180,0 180,0 180,0
Cl 180,0 180,0 180,0 180,0
Met+Cys 714,0 722,0 -34,0 688,0 -25,0 645,0 -15,0
Lys 850,0 - 20,0 840,0 -40,0 800,0 -20,0 750,0
Trp 179,0 -5,0 176,0 -8,0 168,0 -4,0 158,0
Thr 595,0 -14,0 588,0 -28,0 560,0 -14,0 525,0
Ile 672,0 -25,0 664,0 -40,0 632,0 -24,0 593,0 -8,0
Val 765,0 -35,0 756,0 -52,0 720,0 -34,0 675,0 -15,0
Arg 910,0 -47,0 899,0 -67,0 856,0 -45,0 803,0 -23,0
¹ Recommended level according to the management guide for commercial lineages of
semi-heavy hens in 2009; ²Level with positive or negative variation for the lineage
between 2009 and 2014; *Gray spaces represent data without variation, or without
recommendation, in the case of K; **The units of the items are % (Crude Protein),
g/day (Calcium) and mg/day (others).
Overall, there is a reduction in recommended nutritional

levels between 2009 and 2015 for lightweight birds (Table 4). For
the most recent lineage of these layers, there is a considerable
increase in recommended levels, especially in amino acids.
However, there was a reduction of the recommended daily intake
of available P. In the same way, when evaluating the data of semi-
heavy hens, there is a reduction of recommended levels between
2009 and 2014, which was similar for lightweight birds. Probably,
the comparison between most recent lineages to 2014would be
more applied and would have the same effect.
NUTRITIONAL RECOMMENDATIONS FOR

JAPANESE QUAILS
Quail farming in Brazil has shown significant growth in

recent years and the increase in egg production was proportionally
higher, evidencing an important gain in productivity. These results
are attributed to several factors, including precise and efficient
nutrition, aiming to meet the nutritional requirements of birds.
The nutritional requirements change according to the age of
the bird and the purpose, either for meat or egg production. The
birds during their initial phase need nutrients that allow satisfactory
body development for a good productive performance in the
production period.
For years, given the scarcity of data on quail nutritional
requirements, feed formulations were based on requirements of
broiler chickens or laying hens. Nonetheless, it is inaccurate and
does not allow productive results that reflected the true genetic
potential of the quails. Genetics, associated with interspecies
differences, lead to differences in the partitioning of nutrients in the
body, which in turn results in different nutritional requirements.
In the early 1990s, the first research in Brazil began to
investigate and determine the nutritional requirements of
Japanese quails. Research related to the determination of dietary
energy values and digestible amino acids and evaluation of
additives in quails’ diets began to receive more attention from the
researchers. The efforts resulted in two important publications with
tables containing the feed composition and nutritional
requirements of broiler and laying quails: "Tables for European
and Japanese Quails" (Silva & Costa, 2009) and the "Brazilian
Tables for Birds and Swine"(Rostagno et al., 2011). The
publication of these studies allows the formulation of balanced
diets for quails at different stages of development by nutritionists in
both feed processing industry and in academic world.
Although much knowledge is available about the nutrition of
Japanese quail, further information needs to be studied and
elucidated. After 2011, papers that allow a parallel between the

main studies were published to better discuss the nutritional
recommendations at different stages of growth and production of
these birds.
CHANGES FOR JAPANESE QUAILSIN THE

RECOMMENDATIONS OVER TIME
The number of studies on nutritional recommendations for

quails is lower when compared to those for laying hens. In Table
6, it is observed the recommendations for quails in the initial,
growth and laying phases, published by Silva & Costa (2009),
Rostagno et al. (2011) and by different authors between 2011 and
2016.
Protein and amino acids
Comparing the CP recommendations (Table 6) in the

publications between 2009 and 2016, there was no variation in the
requirements. However, the CP requirement is influenced by the
amino acid profile of the diet, since birds do not present CP
requirements itself, but for each one of the essential amino acids
that compose it and for a quantity of nitrogen that is enough for the
synthesis of non-essential amino acids. Hence, it is necessary to
provide a minimum level of protein in the diet; otherwise the
development of the bird may be compromised. Excess protein
may also limit poultry performance, since excess nitrogen in the
form of uric acid needs to be catabolized and this requires extra
energy expenditure.
The use of protein decreases with the advancement of the
age. Silva et al. (2004) reported that the protein retention
efficiency was 40% in the carcass for quails from 1 to 12 days and
23% for quails from 15 to 32 days of age, suggesting that about
60% of the protein intake is lost or oxidized. Quails appear to
retain less protein and energy in the carcass and are less
efficiently than laying hens, reflecting likely differences between

species. In the laying phase, the protein requirement of quails
varies according to many factors such as egg production, quail
weight, balance and availability of amino acids, evaluated criterion
(egg production and weight, feed efficiency, housing conditions)
and answer model. The egg weight is highly dependent on the
daily intake of protein, because laying quails depend on the daily
intake of this nutrient to meet their requirements.
A reduction in Cys, Thr and Arg, those particularly essential
to the development of the birds, was observed in the initial, growth
and laying phases. The requirements in Ile increased in the initial
phase and growth, but the requirements reduced during the laying
phase. These amino acids, besides intrinsically related to the
body's protein synthesis, have important nutritional and metabolic
functions that directly influence the final performance of the
animals. It is worth noting that the final performance of the animal
is not related only to feed intake and feed conversion efficiency.
The performance results depend on several factors and
among them the sanitary challenge is the key. Health-challenged
animals have higher requirements of some amino acids, because
the immune system of birds is required in greater intensity.
Met+Cys, Thr, Ile and Arg have functions directly related to the
immune system.
In recent years, great attention has been given to sanitary
issues and prophylactic measures have been used to prevent
diseases in birds. These measures are taken not only in
commercial farms, but also during experimental trials. Apparently,
this factor contributed to the reduced amino acids requirements.
Table 6 - Variation in nutritional recommendations for Japanese

quails with recommendations from 2009 to 2016, in the
initial, growth and laying phases
1-42 days-old Egg prodution

Nutrient Silva & Rostagno 2011 Silva & 2011
Rostagno
and Costa et al. to Costa to
et al. (2011)
energy (2009) (2011) 2016 (2009) 2016
Protein and amino acids, %
PB 23 22 22.5* 20 20 -
Met+Cys 0.77 0.76 0.74* 0.70 0.88 0.82*
Lys 1.10 1.12 1.25* 1.03 1.08 1.09*
Trp 0.16 0.21 0.21 0.18 0.22 -
Thr 0.86 0.79 0.84 0.67 0.65 0.66*
Ile 0.77 0.80 - 0.87 0.70 0.64
Val 0.77 0.95 0.99* 0.87 0.81 0.89
Arg 1.11 1.19 - 1.38 1.25 1.16
Leu 1.26 1.53 - 1.43 1.62 -
Minerals, %
Ca 0.80 0.90 - 3.05 3.09 3.80
Pd 0.31 0.37 - 0.28 0.32 0.30
K 0.45 - - 0.46 - -
Na 0.24 0.17 - 0.23 0.15 -
Cl 0.15 - - 0.24 - -
EM, kcal/kg 2950 2900 - 2800 2800 -
*Average recommendations.
Crude protein recommendations between 2011-2016:22%, Attia et al. (2012);23.08%, Soares et al.
(2013).
Met+Cys recommendations between 2011-2016: 0.86%, Lima et al. (2016);0.63%, Khosravi et al.
(2016); 0.66%, Scottá et al. (2011); 0.85%, Reis et al. (2011); 0.96%, Sarcinelli et al. (2016).
Lys recommendations between 2011-2016: 1.18%, Lima et al. (2016); 1.30%, Hajkhodadadi et al.
(2013); 1.36%, Mehri et al. (2015); 1.17%, Mehri & Ghazaghi (2014); 1.12%, Ribeiro et al. (2013);
1.07%, Nery et al. (2015).
Trp recommendations between 2011-2016: 0.21%, Mehri et al. (2016).
Thr recommendations between 2011-2016: 0.55%, Mehri et al.(2016); 0.78%, Lima et al. (2013).
Ile recommendations between 2011-2016: 0.64%, Petrucci (2013).
Val recommendations between 2011-2016: 1.04%, Mehri et al.(2016); 0.95%, Cavalcante &
Perazzo (2016); 0.89%, Cavalcante & Perazzo (2016).
Arg recommendations between 2011-2016: 1.16%, Reis et al. (2012).
Ca and available P recommendations between 2011-2016: 3.80 e 0.30%, respectively, Ribeiro et al.
(2016).
Recently, Japanese quails have required higher levels of

Lys, Trp, Val and Leu, for both growing phases (Table 6). An
increase in Lys requirements is directly related to its relationships
with the other amino acids in formulations based on the ideal
protein, since it is used as the reference amino acid. Increases in
Lys requirements would theoretically reflect in increased levels of
other amino acids, but not all amino acids behaved in this way. As
can be seen, only Trp, Val and Leu showed such behavior.
Consequently, the relationships between amino acids and Lys are
changing in recent years and have not followed the relations that
Rostagno et al. (2011) suggested.
Besides a reference amino acid, Lys is considered
physiologically essential for maintenance, growth and production.
It acts mainly in muscle protein synthesis, bone formation (Vieira,
1999) and carnitine synthesis, which acts in the transport of fatty
acids for β-oxidation in mitochondria. In corn and soybean meal-
based diets, Lys is the second limiting amino acid for Japanese
quails in the growing and breeding periods (Cavalcante, 2013).
Trp is considered the third limiting amino acid in corn and soybean
meal-based diets for laying hens. This amino acid is part of the
structure of all proteins and it is precursor to the synthesis of
serotonin and melatonin (Corzo et al., 2005). In metabolism, Trp is
converted to niacin (vitamin B3), which is a precursor of two
coenzymes that participate in almost all oxidation reactions in the
body: Nicotinamide adenine dinucleotide (NAD+) and Nicotinamide
adenine dinucleotide phosphate (NADP+).
Valine and Leu are part of the branched chain amino acid
group with Ile and they compose35% of the proteins in the muscle
tissue of birds, as well as participating in the feather formation.
These amino acids are highly hydrophobic and are generally found
within proteins, participating in the three-dimensional structure of
proteins (Lehninger, 1991). Although it is essential for muscle
growth and feathering, the equilibrium in the supply of these amino
acids is fundamental for good performance, because these three
amino acids compete for the same sites of absorption and for the
same enzymes in the metabolism. Diets with excess Leu increase

the catabolism of Val and Ile because their alpha-ketoacids
intensify the dehydrogenase activity of keto acids of branched-
chain amino acids, which increases the degradation of Val and Ile.
Minerals
Minerals have several functions in the animal organism.

They are required for skeletal formation, as components of several
compounds with particular functions within the body, as enzyme
cofactors and for the maintenance of osmotic balance within the
body of the bird. Among the minerals, more attention is given to
Ca and P in the formulation of diets for quails.
Japanese quails have become more demanding in recent
years for Ca and P (Table 6). In the growing and breeding phases,
Ca guarantees proper bone mineralization. Birds with Ca and P
deficiency have growth retardation, decreased intake and bone
fragility during the development phase. Approximately 99% of the
body's calcium is deposited in bones (Nunes et al., 2006). During
the growing and breeding phases, Ca demand is relatively low
because there is no eggshell formation. However, from sexual
maturity, at around 42 days of age, this demand changes
drastically and the bird requires a calcium intake approximately
four times higher than the demanded in the previous period.
In egg production, it is of fundamental importance to verify
dietary Ca and P levels because they are directly related to the
egg quality. For adult birds, Ca is used for eggshell formation, but
its excess negatively interferes in the availability of other minerals
such as P, magnesium (Mg), manganese (Mn) and zinc (Zn).
During the calcification period, the Ca to be deposited in the
eggshell has two origins: dietary and bone. Even with proper
dietary intake, about 30 to 40% of the Ca deposited in the eggshell
comes from the bone marrow. However, the use of skeletal Ca
can be increased because of the low intake. The greater the Ca
dependence on the skeleton, the smaller the amount deposited in
the eggshell, resulting in eggs with thin shells and little resistance
to breaking. Therefore, the dietary Ca supply to meet poultry

demands not only targets egg production, but also a good eggshell
quality.
Phosphorus, as well as Ca, also participates in bone
mineralization and eggshell formation, but in lesser amounts.
However, it is necessary to balance the supply of these two
minerals, since the excess of one interferes in the absorption and
use of the other (Lopes, 2011). In addition to participating in bone
formation, the P participates in the constitution of the main
metabolic energy carrier molecule, the ATP molecule.
Sodium, Cl and potassium (K) are important elements to
maintain osmotic pressure and electrolyte balance within normal
values (Murakami & Franco, 2004). These electrolytes in body
fluids are specifically involved in water metabolism, nutrient
absorption and transmission of nerve impulses at the cellular level.
Currently, micro minerals, also called trace minerals such as Zn,
Mn and selenium (Se), have been studied in bird nutrition. Interest
in these minerals is related to their functions, since they act as
components of protein structures or as cofactors, acting in
allosteric alteration or modulation in the tertiary structure of
enzymes, making them active or inactive.
METABOLIZABLE ENERGY
The requirements for metabolizable energy did not show

differences between the years, but recent studies seeking to
update energy requirements for Japanese quails (Table 6) are
scarce. Energy level is one of the most important components in
the feed formulation for quails, considering that birds initially
increase their intake to meet the energy needs. In a breeding
system where feed is used ad libitum, feed intake is regulated by
the energy density of the feed and by the requirement of the birds.
Therefore, the precise knowledge about the feed energy is
necessary to provide the adequate balance of the diets. However,
the effectiveness of the feed formulation method is dependent on

the accuracy with which feed energy is determined.
Feed intake is regulated by dietary energy density and
nutritional requirements, affecting egg performance and quality
(Moura et al., 2010). Therefore, according to Bertechini (2012), all
nutrients must be related to the dietary energy content. Excess
dietary energy in the form of carbohydrates, lipids and proteins
(imbalance in the amino acid profile) causes a reduction in the
voluntary intake (Baião & Lara, 2005) and deposition of fat in the
carcass. According to Bertechini (2012), excess fat in the liver and
ovary may occur due to energy imbalance in the diets of
commercial laying hens, promoting a reduction in egg production.
FINAL CONSIDERATIONS
Considering the above, poultry are becoming smaller and

even more demanding because they efficiently convert feed into
an extremely rich and functional food, the egg. This is an important
and fundamental advance for the productive sector. In this context,
the demand to update the nutritional needs of laying hens is
constant, not only to meet demand and to produce well or
satisfactorily, but to meet a genetic, physiological, nutritional,
environmental, economic and sustainable demand.
Japanese quail farming has become more expressive and
there is a need for more research that determines the nutritional
requirements for these birds, always seeking the productive
efficiency to increase farmers’ profitability and to obtain a product
of great quality for the consumer.
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INORGANIC AND ORGANIC TRACE MINERALS
NUTRITION OF BROILERS
Ramon D. Malheiros and Peter R. Ferket
Prestage Department of Poultry Science, NC State University, Raleigh, NC,
USA, 27698-7608
ABSTRACT
This experiment was conducted to evaluate the effect of

organic and inorganic trace minerals in broilers diets. A total of
1872 1 day-old male broiler chicks (Ross 780) were placed in a
broiler house with 72 pens. Two sources of supplemental trace
minerals: inorganic and organic and levels of trace minerals: 12.5,
25.0, 37.5 and 50.0%, plus the positive control with 100%
supplementation of inorganic mineral were used in a 2 x 4 + 1
factorial arrangement in 8 randomized complete blocks
considering the positioning inside the broiler house with 26 birds
per experimental unit. The results of this experiment demonstrate
that organic trace mineral resulted in better feed conversion,
viability, and superior growth performance over inorganic trace
minerals, especially at low dietary inclusion levels. The organic
mineral supplement increased ash, P, Ca, and Mn content of tibia
at the lowest level of trace mineral supplementation. Litter
concentration of Zn, Cu, and Mn decreased as the dietary level of
trace mineral decreased. Regardless of supplementation level, the
organic form resulted in lower pH of meat breast than the
inorganic mineral source. Moreover, dietary supplementation of
the organic trace mineral source resulted in decreased drip loss of
meat breast during 7 day storage. Finally, footpad lesion score
was significantly reduced by the supplementation of organic trace
minerals. Total replacement of inorganic trace minerals by low
levels of organic forms is beneficial for growth performance,
welfare, and meat quality of broilers.
124 - IV International Symposium on Nutritional Requirements of Poultry and swine
INTRODUCTION
Minerals, such as copper (Cu), iron (Fe), manganese (Mn),

and zinc (Zn) and are essential for the growth of chickens and are
involved in various physiological processes. They participate in
nearly all metabolic pathways of the animal body, and they have
physiological functions that are essential to life, such as
reproduction, growth, immune system, bone formation and energy
metabolism (Dieck et al., 2003; Bao et al., 2007; Dibner et al.,
2007).
With the intensification of agricultural production, many
foods used as ingredients in diet formulation experienced a
decrease in the trace minerals concentration (Graham et al., 1999;
Garvin et al., 2006). Although the National Research Council
(NRC, 1994) provides requirement values of trace minerals values
for poultry, but many of these values were determined decades
ago on slower growing strains or the values were simply
estimated. The requirements of trace minerals for broilers refer to
the same levels recommended by NRC in the early 1990’s, with
some being based on data from the 1950’s (Bao et al., 2007). With
this, nutritionists often utilize higher levels of minerals, frequently
based on their own practical knowledge (Leeson, 2008).
Practically, increasing the margin of safety for mineral
supplementation well above actual nutritional requirements results
in excessive mineral excretion. Obviously, this is not only wasteful,
but also has adverse consequences on environmental
sustainability. The concern about mineral buildup in the
environment, especially Zn and Cu, leads to questions about
mineral source bioavailability and how to reduce the levels of
mineral supplementation in the diets without compromising the
nutrition of animals and the quality of their products. The use of
organic minerals has been suggested as a solution to this
problem, based on the assumption that the mineral complexed in
the form of chelates have an increased bioavailability in
comparison to inorganic sources, such as sulfates, carbonates
and oxides (Ji et al., 2006). Nutritional bioavailability of organically

chelated minerals may be due to reduced competitive binding of
mineral elements with fiber, phytate, Ca or P during digestion
(Brooks et al., 2012). Thus, trace minerals supplemented to the
diet at lower concentrations as organic chelates than inorganic salt
forms, without adverse effect on productive performance or
excessive excretion into the environment (Bao et al., 2007).
There are different forms of organic minerals available for
use in animal nutrition applications, and the bioavailability of these
organic minerals these have been investigated by several
researchers. Among these organic trace mineral complexes or
chelates, the proteinate trace mineral chelate has shown
nutritional benefits (Bao et al., 2007; Nollet et al., 2007; 2008;
Abdallah et al., 2009). Proteinate trace minerals generally
comprise of proteins, peptides and amino acids, and they are
formed by the reaction of soluble inorganic salts with mineral
hydrolyzed vegetable protein (Nollet et al., 2007). The reaction of
the mineral with the hydrolysate results in the formation of
complexes containing chelated metallic ions. Studies have
demonstrated that the trace mineral bioavailability varies
substantially among different sources; trace minerals linked to an
organic molecule (organic minerals) have a bioavailability that is
substantially better than inorganic forms because they reduce
antagonistic reactions with other feed components in the
gastrointestinal tract (Richards et al., 2010; Manangi et al., 2012).
Perhaps because of differences in mucosal transport, the use and
the deposition of these trace minerals are differentiated in the
target tissue and organs; however, this mechanism has not been
elucidated. It was hypothesized that the use of the most
bioavailable trace minerals as organic complexes allows
nutritionists to decrease the mineral content in the diet without
compromising nutritional requirements (Manangi et al., 2012). The
purpose of the present study was to compare the efficacies low
dietary supplementation levels of inorganic and organic trace
mineral sources (supplemental zinc, manganese, copper, and iron
at 12.5, 25.0, 37.5 and 50.0% of the industry standard) on growth

performance, minerals concentrations in tissue, mineral excretion,
quality and lipid oxidation of meat, and relative enzyme activities.
MATERIAL AND METHODS
A total of 1872 day-old male broiler chicks (Ross 780) were

randomly assigned among the dietary treatment groups. The
experimental diets were based on corn and soybean meal,
formulated to meet or exceed the NRC 1994 requirements except
for Cu, Mn, Zn and Fe. All diets had similar nutrient composition
and the same concentrations of Zn, Mn, Cu, and Se, the difference
being in the relative amounts of organic and inorganic (sulfate)
forms. The experiment was designed as a 2 X 4 factorial
arrangements of treatments: two sources of supplemental trace
minerals (inorganic and organic), and 4 levels of trace minerals
(12.5, 25.0, 37.5 and 50.0% of industry standard level), plus the
positive control with 100% supplementation of inorganic mineral.
Each dietary treatment was replicated in 8 pens containing 26
birds per pen. Birds had ad libitum access to water and feed
during the study. Birds were weighed at1, 14, 32 and 48 days of
age in order to determine weight gain. At 32d and 48d, two birds
per experimental unit were killed by cervical dislocation and the
right legs were collected. Breast samples from these same birds
were collected to determine drip loss over time, muscle color, and
pH. Another breast portion was taken from each sampled bird,
properly identified, and stored frozen at -20o C prior to the
determination of lipid oxidation. Poultry litter samples were
collected to quantify the concentration of minerals (Cu, Zn, Mn and
Fe) at 32 and 48 days. Breast and heart samples were collected,
identified, frozen on liquid nitrogen, and stored at - 70° C for later
analysis of superoxide dismutase activity (SOD).
Statistical Analysis: All data were analyzed as a
completely randomized block design by ANOVA using a 2 x 4
factorial arrangement of mineral source and supplemental mineral
level as main effects, and their interaction effects. Treatment main
effects and interaction effects were declared significant at

probability less than 5%. Data were subjected to ANOVA using the
GLM procedure of SAS system (SAS Institute, 2003). After
ANOVA, all treatments were compared with the positive control
(100% inorganic mineral) by Dunnet test, probability was less 5%.
RESULTS
Performance: Organic mineral improved the BW of birds at

48 d, although this significant effect was not observedat14 and
21d. Birds fed the organic mineral treatments had better feed
conversion and lower mortality rate than those birds that received
the inorganic forms of trace minerals. At32 d of age, the 12.5 and
25.0% dietary inclusion level of trace minerals in the organic form
resulted in better in feed conversion than offered at the same
levels as the inorganic form. Birds fed 12.5 and 37.5% level of
trace minerals had lower viability than those who received 100%
level of inorganic mineral. However, birds fed organic trace
mineral diets had a higher production efficiency index than those
fed the inorganic form.
Mineral Concentration: Tibia: At 32d, the tibia of the birds
who received 37.5% of inorganic mineral had a lower percentage
of ash as compared to those who received the same amount of
organic mineral. There was no significant effect on the
concentration of Cu and Zn as well as percentage of Ca and P in
the chicken’s tibia at 32 d. At48 d, birds fed the organic trace
minerals had higher tibia ash content than those fed the inorganic
for of trace minerals. Birds fed organic minerals had higher Ca and
P content of tibia than those who received the inorganic form of
the trace minerals.
Liver: There was no significant difference in the percentage
of ash, as well as the concentrations of Mn and Zn in the liver of
the birds at 32 days of age.
Litter: At 32 and 48 d, there was no significant difference
between treatments for the results of ash and Fe in the litter.
However, there was a direct correlation between the amount of
ingested minerals and the concentrations of Cu, Mn and Zn in the

litter, in the extent to which increased mineral in diet increased the
percentage concentration of the mineral the litter. At 48 days, it
was observe that birds fed increasing levels of inorganic or organic
minerals excreted smaller amount of Cu, Mn and Zn as compared
to the positive control group (100% mineral inorganic).
Meat quality: Color: No significant mineral source or level
effects was observed on meat color or meat color change during
storage except at 32 d: birds in 25.0% level of inorganic mineral
group had higher lightness values as compared to those who
consumed 25.0% of organic mineral source. At 48 d, birds that
consumed 25.0% of inorganic mineral showed higher yellowness
values as compared to those of control group.
pH and drip loss: There was no significant difference in
breast meat pH at32 d or drip loss at48 d. At 48d, the breast pH of
birds fed the organic minerals was greater than those fed the
inorganic mineral.
Lipid oxidation (MDA) and superoxide dismutase
(SOD): There was no significant difference for the MDA
concentration in muscle breast of birds at32 and 48 d, as well as
the concentration of SOD in the heart of the birds at 48 d.
Bone strength: There was no significant effect between
the results of treatments for bone strength of broiler tibia.
CONCLUSION
In conclusion, the results of this experiment demonstrate

that organic trace mineral resulted in better feed conversion,
viability, and superior growth performance over inorganic trace
minerals, especially at low dietary inclusion levels. Dietary
supplementation of organic minerals increased tibia ash, P, Ca,
and Mn content at the lower levels of trace mineral
supplementation. The lower dietary supplementation levels of
trace mineral reduced the Zn, Cu and Mn concentration of litter.
Beast meat quality was also improved by organic trace mineral
supplementation, as indicated by lower pH, and reduced drip loss.
Moreover, animal welfare and paw quality was improved as food

pad lesion score was reduced by dietary supplementation of
organic trace minerals.
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RECOMMENDED VITAMIN LEVELS FOR HIGH
PERFORMANCE BROILERS AND LAYERS
Gilberto Litta
Animal Science & Advocacy Manager
DSM Nutritional Products, Animal Nutrition and Health,
Via G. Di Vittorio 20090 Segrate (Italy)
gilberto.litta@dsm.com
ABSTRACT
Poultry industry worldwide is committed to achieve best

performance, feed utilization, welfare and health of birds. Optimum
nutrition with both macro and micronutrients is pivotal for ensuring
health and performance. Vitamins are micronutrients added to
poultry diets in small amount but having a big impact. Nowadays
clinical vitamin deficiency symptoms are rarely observed, mostly in
advanced farming systems, but sub-clinical vitamin deficiencies
often penalize animal’s performance and health. In this paper, we
aimed to provide an overview about the chemical characteristics,
the metabolism, the functions and an overview about the main
results obtained in using vitamins in broilers and poultry egg
nutrition.
INTRODUCTION
Optimum nutrition occurs only when poultry receive the

correct mix of macro- and micronutrients in the feed for efficiently
utilize those nutrients for its growth, health, reproduction -
including embryonic development, hatchability and chick viability -
and survival.
Vitamins are micronutrients, essential for life of man and
animals to enable the animal to efficiently utilize all other nutrients:
they play a key role in the metabolism. As most vitamins cannot be
synthesized by poultry, they must be obtained from the feed. If
vitamins are absent from the diet or improperly absorbed or

utilized, specific diseases or deficiency disorders occurs.
Vitamins are recognized by two properties: (1) the daily
requirement for each vitamin is few micrograms or milligrams; (2)
vitamins are organic compounds, not building substances,
exercising catalytic functions: they facilitate both synthesis and
degradation of the nutrients, thereby controlling metabolism.
Classically, vitamins are classified based on their solubility:
(1) the fat-soluble group including the vitamins A, D3, E and K and
(2) the water-soluble vitamins including the B complex (B1, B2, B6,
B12, niacin, pantothenic acid, folic acid and biotin) and vitamin C.
The fat-soluble vitamins are absorbed by mechanisms
similar to those involved in fat absorption while the uptake of
water-soluble vitamins occurs usually via simple diffusion.
Fat-soluble vitamins may be deposited in the animal body,
while water-soluble vitamins are not stored and excesses are
rapidly excreted: their regular uptake is therefore even more
essential for securing adequate vitamin availability.
The water-soluble vitamins of the B-complex are, most of
them, coenzymes that can accelerate various metabolic
processes. Vitamin B1, vitamin B2, vitamin B6, niacin, pantothenic
acid and biotin are all involved in the energy metabolism, while
vitamin B12 and folic acid exert their activity on growth and cell
maintenance. It is however important to consider that those B-
vitamins, related to a particular metabolic effect, are interacting
with each other, which makes it difficult to determine individual
requirements for each vitamin of the B-group.
Thanks to continuous improvement in genetics, today’s
poultry breeds are very productive: hence, also their vitamin
nutrition needs particular attention in order to allow the birds to
perform up to their genetic potential. Today the supplementation
of vitamins to animal feeds is not disputed: questions are about
the supplementation levels necessary to profitably achieve
optimum health and performance under production conditions.
VITAMIN DEFICIENCIES
In general, vitamin deficiencies generate loss of appetite,

mild to severe growth retardation and impaired feed conversion,
which result in bad performance. Main deficiency symptoms will be
described later in this text for each individual vitamin. Clinical
vitamin deficiencies although rare, and sometimes associated with
stressful situations, can still occur in poultry under production
conditions. Some cases have occurred, even recently, due to
complete absence of supplemental vitamin in the feed (Cortes et
al., 2006).
An authoritative source of information about the minimum
vitamin levels necessary to avoid clinical deficiency symptoms is
the nutrient requirements of the National Research Council (NRC,
1994). Many of the references are outdated, being 20 to 40 years
old, and many studies were carried out under perfect husbandry
conditions (and it is known that vitamin requirements are lower in
absence of the stressors occurring under commercial conditions).
To optimally exploit the continuous improvement in
genetics, to strengthen the resistance against various infectious
diseases, to improve welfare and to optimize meat and egg
quality, the vitamin nutrition of poultry must be adapted to today’s
husbandry and management conditions. Therefore, breeding
companies, local country organizations such as the German Trade
Association (AWT, 2002), Universities (Universidad ederal de
Vi osa, 2011 and vitamin suppliers , 2016 have published,
and most of them regularly update, own vitamin supplementation
guidelines, which are based on experience from industry practice
as well. These supplementation levels generally exceed the
minimum requirements provided by NRC, since aiming at
achieving optimum health and performance of domestic animals in
the most cost-effective way.
GENETIC DEVELOPMENT OF POULTRY BREEDS
A considerable part of the classical vitamin requirement

studies, which have been taken as a basis for the
recommendations of the NRC (1994), date back to the years
1960-1980. The poultry breeds, both meat-type and laying, which
were used decades ago have nothing in common with current
modern genotypes (Havenstein et al., 2003). Since the dietary
inclusion levels of vitamins are expressed in units of feed, the
absolute uptake of vitamins for an equivalent muscle or egg
production has steadily decreased accordingly to improved FCR.
Whether a constant adjustment is necessary for the growing
genetic potential of modern breeds has been addressed in several
studies.
Fat Soluble Vitamins
Vitamin A (Retinol)
Characteristics and Properties of Vitamin A
Vitamin A (retinol), a fat-soluble vitamin, is an alcohol, but it

occurs in nature mainly as a fatty acid ester. It is found only in
animal origin foods and mostly in fish like cod and tuna. Out of
various isomers, only the all-trans-vitamin A conveys the full
biological activity. Beta-carotene, the pro-vitamin A, occurs
predominantly in the vegetable kingdom: the molecule is cleaved
in the intestinal epithelium releasing retinol, which is absorbed, like
vitamin A retinol, in conjunction with low-density lipoproteins
(LDLs). Light, oxygen and acids rapidly destroy both the alcohol
and the ester forms. Vitamin A is usually commercialized as retinyl
acetate or retinyl palmitate in oily solutions, in stabilized powders
or in aqueous emulsions. An International Unit (IU) corresponds to
the activity of 0.344 mcg of pure crystalline vitamin A acetate.
Vitamin A is essential for normal vision, since it is part of the

visual pigment rhodopsin, it supports the development and
maintenance of the integrity of epithelia, mucous membranes and
skeleton and it supports the immune response. In farm animals,
vitamin A is important for normal feed intake, growth and
performance. Depending on the poultry category, the age and the
period of the life cycle, the general recommendations reside
between 8’000 and 15’000 IU vitamin A/kg of feed.
Vitamin A deficiency induces loss of appetite, growth
retardation/inhibition and eventually can result in death. The birds
develop a dry and scaly skin, a rough plumage, hyperkeratosis of
the cornea and metaplasia of respiratory epithelia, which
increases the risk of infections. Keratinization of epithelia in the
digestive tract depresses gland activities and results in poor
absorption of nutrients. In laying hens, a marked decline in egg
production can occur due to severe degeneration of the ovary.
Vitamin A in Broilers
Yet since all fat-soluble vitamins A, D, E and K are

absorbed via the same channels in the intestine, interactions
among them can occur. High levels of vitamin A - up to 50’000
IU/kg - were found to decrease plasma levels of vitamin E, while
high vitamin E levels had no effect on circulating retinol (Frigg and
Broz, 1984). Likewise, high dietary levels of vitamins A and E
negatively affected the utilization of vitamin D3, but only when D3
was present at marginal levels (Aburto and Britton, 1998a). These
findings indicate that it is critical to provide fat-soluble vitamins in
correct ratios and that excessive or defective level of one of them
can have an impact on the others. The effect of vitamin A on the
small intestine was examined in vitamin-A-deficient meat-type
chickens (Uni et al., 2000). Vitamin A deficiency caused
hyperproliferation of enterocytes, a decrease in the number of
goblet cells, and other effects suggesting that the absence of
vitamin A interferes with the normal growth rate in chickens

because it influences functionality of the small intestine.
Amongst micronutrients, vitamins A, D, and E were
demonstrated to have a direct modulating activity on the immune
system (Klasing, 1998). Leutskaya and Fais (1977) have
demonstrated that the antibody content in chickens depended on
the dose of vitamin A in the diet. In addition, T-lymphocyte
proliferative responses were decreased at low vitamin A intakes
and enhanced at the high vitamin A intake (Sklan et al., 1989).
Insufficient, but also excessive, vitamin A led to increased
susceptibility of chicks to E. coli infection and depressed immune
responses (Friedman et al., 1991). In broiler chickens challenged
with Newcastle disease virus (NDV), both humoral and cellular
immune responses were modulated by dietary vitamin A (Lessard
et al., 1997). Broiler chicks challenged with coccidiosis revealed
that insufficient supply of vitamin A compromised local immune
defenses as reflected in lymphocyte profiles, oocyst shedding and
interferon-gamma levels (Dalloul et al., 2002). Vitamin A showed
also important protective effect against heat stress. When broilers,
supplemented with 15’000 IU vitamin A/kg feed, in combination
with zinc, were reared at 34 °C live weight gain, feed efficiency
and carcass yield were significantly improved over a control with
lower A-levels, indicating that zinc and vitamin A have similar
effects in preventing heat-stress-related depression in
performance of broiler chickens (Kucuk et al., 2003).
Vitamin A in Laying Hens and Breeders
In a series of experiments, Richter et al. (1990) established

that a minimum of 5’000 IU vitamin A per kg are necessary to
allow adequate feed intake and laying performance during the
production period. Higher levels did not provide more benefits,
probably due to the limited genetic potential of those hens at the
time. Squires and Naber (1993) showed that Vitamin A levels of
egg yolk were not related to dietary level and therefore are not
useful to predict future vitamin A deficiency in laying hens. Vitamin

A deficiency in laying hens was found to lower overall egg
production and to induce the appearance of atretic follicles,
containing moderate to severe hemorrhages (Bermudez et al.,
1993). In heat-stressed commercial layers 12,000 IU/kg of vitamin
A improved feed intake, laying performance and egg weight of
hens (Lin et al., 2002). A good vitamin A supply is also essential
for pullets (Beynen et al., 1989).
Finally, adequate vitamin A is beneficial on the various
parameters of the immune responses. Lin et al., 2002 observed
that 12,000 IU/kg increased the antibody titer against NDV virus of
heat-stressed hens.
Vitamin D3 (Cholecalciferol)
Characteristics and Properties of Vitamin D3
Poultry can only utilize vitamin D3 (cholecalciferol) not D2.

Rich sources of vitamin D3 are liver and viscera of fish. Vitamin D3
can be endogenously produced by UV irradiation of 7-
dehydrocholesterol, but since most of the skin of poultry is covered
with feathers, dietary Vitamin D3 is the most reliable source.
Vitamin D3 is a colorless crystals, soluble in alcohol and other
organic solvents, but less soluble in vegetable oils, sensitive to
oxygen and acids. One International Unit (IU) is equal to 0.025
mcg cholecalciferol (or 1 mcg=40 IU).
Cholecalciferol is absorbed from the intestinal tract with fats
with the intervention of bile salts, via the lymphatic system is
transported to the liver and is finally deposited in adipose tissues.
In the liver cholecalciferol is converted to 25-hydroxy-
cholecalciferol (25OHD3) and then further hydroxylated in the
kidney to 1,25-dihydroxy-cholecalciferol (1,25OH2D3). The last
transformation step to the active hormonal form of vitamin D 3 is
under the control of the parathyroid hormone. Vitamin D3 regulates
the homeostasis of calcium and phosphorous, i.e. it increases the
absorption of calcium and phosphorous from the small intestine as

well as the re-absorption of these minerals in the renal tubules and
increases the uptake of minerals by the bones. A dietary
supplementation of 3’000 to 5’000 IU/kg feed is generally
considered adequate for the various poultry categories.
In all higher animals, the best-known and most dramatic
endpoint of vitamin D3 undersupply are rickets and osteomalacia.
Clinical deficiency signs in poultry are inhibition of growth, loss of
weight, reduced or lost appetite and high mortality. The epiphyses
are enlarged; the bones of the extremities, of the vertebral column
and the cranium are deformed and brittle. The beak of rachitic
chicks is soft and pliant. Animals move stiffly and hesitantly due to
occurrence of lameness and muscular weakness. Egg production
decreases and eggs with thin shells are laid.
Vitamin D3 in Broilers
Vitamin D3 is always supplemented to commercial poultry

feeds. While NRC (1994) still recommends a level of 200 IU/kg,
practical supplementation has reached levels of 2’500 to 5’000
IU/kg. A comparative study with supplementation levels of 2’500
IU/kg versus 200 IU/kg confirmed that broilers on the higher
vitamin D3 level consumed more feed, gained more weight and
had a higher percent tibia ash (Ledoux et al., 2004) and a dietary
supplementation with 10’000 IU/kg feed significantly improved
growth, enhanced tibia breaking strength and reduced the
incidence of tibial dyschondroplasia (TD) in comparison to 5’000
IU/kg (Whitehead et al., 2004), indicating that modern broiler
genotypes have increased requirements of the micro-nutrients
supply.
As said high supplementation levels of vitamins A and E
were found to negatively affect the utilization of vitamin D3 (Aburto
and Britton, 1998a) and to adversely affect bone ash and plasma
calcium in broilers (Aburto and Britton, 1998b) indicating that a
proper ratios of vitamins A, D3 and E in needed.
By feeding 25OHD3 its plasma level, the indicator of vitamin

D nutrition, is always higher compared to any Vitamin D3 in feed,
hence ensuring an optimal vitamin D nutrition even in case of
malabsorption (Rebel and Weber, 2009).
Tibial dyschondroplasia (TD) represents an important
welfare issue for the poultry industry. Although vitamin D3 as such
was described not to be effective, several vitamin D 3 metabolites
were found to prevent the development of TD in broilers (Edwards,
1990). A number of studies have been conducted by supplying
25OHD3, the only globally authorized and commercially available
metabolite. Mireles et al. (1996) conducted a series of experiments
showing that replacement of portions of vitamin D3 with 25OHD3
significantly reduced the incidence of TD, which in turn tended to
improve weight gain. Two other studies (Mireles and Sun, 1997)
showed that birds receiving 25OHD3 had significantly better weight
gain and lower incidence of TD than birds receiving control feed.
With a TD inducing diet the incidence of TD in broilers was
decreased when 70 mcg/kg 25OHD3 was supplemented (Ledwaba
and Roberson, 2003). Wideman et al. (2015) showed that
25OHD3 has a potential to attenuate outbreaks of late lameness
caused by BCO in commercial broiler flocks. Finally, Saunders-
Blades and Korver (2006) showed that improvement of bone
mineralization by inclusion of 25OHD3 could reduce incidence of
Black Bone, a defect caused by blood leakage and bone
discoloration, and improve the consumer acceptance of poultry
meat.
Yarger et al. (1995) in a series of 10 trials conducted on
more than 36,000 broilers showed for all 10 studies positive and
significative effects of 25OHD3 on body weight gain, FCR and
breast meat yield, with a significant dose-response relationship:
maximal effects have been observed at 50 to 70 mcg/kg feed.
Similar effects have been reported by Santos and Soto-Salanova
(2005) and Bray et al. (2012) working on birds vaccinated against
coccidiosis. The underlying molecular mechanisms for body
weight gain and more specifically breast meat yield improvement
was unknown since recent years when some specific studies have
been carried out. The trial carried out by Hutton et al. (2014)
suggest that by feeding 25OHD3 can stimulate satellite cells
activity, the cells driving muscle hypertrophy, in the predominantly
fast-twitch Pectoralis major muscle and provide evidence toward
understanding the mechanism behind previously observed
increases in breast meat yield in 25OHD3-fed commercial broiler
chickens. Similar results have been reported by Berri et al. (2013)
and confirmed by Vignale et al. (2015).
The discovery of Vitamin D Receptors (VDRs) in cells other
those involved in mineral and bone homeostasis strongly indicated
a more diverse role of vitamin D than originally accepted. VDRs
and the enzyme 1alpha hydroxylase have been discovered in
almost all immune cells including T and B cells, neutrophils,
macrophages and dendritic cells (Provvedini et al., 1983;
Shanmugasundaram and Selvaraj, 2012). In recent years some
researches have demonstrated that 25-OH-D3 in poultry diets
plays a role in modulating immune response (Mireles A., 1997;
Mireles et al. 1999). Morris et al. (2014) showed that 25OHD3, but
not Vitamin D3, improved immune response in LPS challenged
birds and that it induce a specific immune response by reducing
pro-inflammatory cytokines and stimulating nitric oxide production
and hence improving immune response.
Vitamin D3 in Laying Hens and Breeders
In adult laying hens, vitamin D3 deficiency produces

osteomalacia with decreased concentration of calcium (Ca) and
phosphorus (P) in the bone matrix, reduced egg production and
egg weight and increases the occurrence of thin shells, cracks and
deformities. Eggshell pimpling is inversely related to the level of
cholecalciferol in the diet (Goodson-Williams et al., 1986) and 4
weeks on a vitamin D3-deficient diet reduce the thickness of the
shell and to induce production of numerous thin-shelled and soft-
shelled eggs. The outer layers of the shell are reduced or absent
but the inner mammillary layer is always present, suggesting that

the hens stop laying before Ca concentrations in blood become
too low for the formation of the mammillary knobs. In laying hens
kept under commercial conditions clinical signs of vitamin D3
deficiency such as cage-layer-fatigue or thin-shelled eggs can
frequently be observed, indicating insufficient utilization of the
dietary vitamin D3. Supplementing 25OHD3 has been proposed to
counteract such problems. García et al. (2001) tested 25OHD3 in
layer diets, on top of a basal supply of vitamin D 3. At similar
performance, eggshell thickness was better in the 25OHD 3
treatment, demonstrating that this metabolite can improve eggshell
quality without any adverse effects. In another experiment, various
combinations of vitamin D3 and 25OHD3 were evaluated in laying
hens (Koreleski and Swiatkiewicz, 2005). At 66 and 70 weeks of
age, partial or complete substitution of cholecalciferol with
25OHD3 increased eggshell breaking strength. Practical type
studies have shown that replacing vitamin D3 partly by 25-OH-D3
improved laying performance, decreased the number of broken
eggs and increased egg weight, egg mass as well as the number
of extra-large eggs (Soto-Solanova and Hernandez, 2004). In
addition, egg yolk susceptibility to oxidation was decreased and
the weight loss in eggs during storage was numerically reduced.
Likewise, under commercial conditions a numerical improvement
of laying performance with 25OHD3 supplementation could be
observed (Soto-Solanova and Schliffka, 2007). The effect of
25OHD3 administered to breeders is transferred via yolk and
embryo to day-old chicks.
In breeders, vitamin D3 is necessary for embryo hatchability
and viability. Hens maintained on 25OHD3 produce normal
embryos whereas when maintained on 1,25OH2D3 as their sole
source of vitamin D produce eggs which appear normal but which
produce embryos having a defective upper mandible and which
die at 18 to 19 days of embryonic life. It therefore appears that
1,25OH2D3 is not transferred from hen to egg in sufficient amounts
to support embryonic development and that vitamin D or 25OHD3
or both, are necessary for normal chick embryo development

(Sunde et al., 1978). Saunders-Blades and Korver (2008)
investigated the effects of maternal and dietary 25OHD3 on chick
immunity and growth. Embryonic mortality was lower and body
weight was improved in the 25OHD3 maternal treatment (69
mcg/kg feed) than in the vitamin D3 treatment (2,760 IU/kg). In
vitro chick leukocyte E. coli killing capability was improved with
maternal 25OHD3. The conclusion was that both maternal and
dietary 25OHD3 improved aspects of broiler production and
immune function.
Vitamin E (alpha-Tocopherol)
Characteristics and Properties of Vitamin E
Vitamin E is found in cereal germs, most oilseeds and in

leafy vegetables as well as in animal organs. Vitamin E occurs in
nature as a series of compounds called tocopherols with alpha-
tocopherol being the most effective for poultry. Alpha-tocopherol is
a bright yellow viscous oil, which is sensitive to oxygen. Therefore,
it is commonly commercialized in the form of alpha-tocopheryl
acetate, which is stable, but can easily be hydrolyzed before
absorption in the intestine. One International Unit (IU) is defined as
1 mg dl-alpha-tocopheryl acetate, which is equivalent to 0.909 mg
dl-alpha-tocopherol.
The absorption of vitamin E is linked to fat absorption,
facilitated by the presence of bile salt. There is a negative
interaction with the absorption of vitamin A (high A levels depress
E absorption). Transport from the intestine to the systemic
circulation runs via chylomicrons of the lymph. In plasma, vitamin
E is bound to lipoproteins and the main storage of vitamin E
occurs in the liver.
Alpha-tocopherol is nature’s most powerful fat-soluble
antioxidant and as such is important to protect the phospholipids
of cellular and sub-cellular membranes from destruction by lipid
oxidation and accordingly to maintain the morphological integrity

and functionality of cells and tissues of the organism. Furthermore,
as an essential micronutrient vitamin E supports optimum
performance and reproduction of farm animals. In poultry, vitamin
E protects the ovarian follicles from oxidative damage and
facilitates the release of vitellogenin, a precursor of the yolk from
the liver, and thus has an important function in the process of egg
production. Due to its modulatory effect on the immune system,
(activation of macrophages, production of antibodies) alpha-
tocopherol has proven effective for the prevention and resistance
against various diseases. Normal supplementation levels to meet
physiological requirements in poultry range between 20 and 100
mg/kg. For special applications such as modulating the immune
response or improving meat quality, dosages up to 250 mg/kg are
needed.
Signs of clinical vitamin E deficiency include muscular
myopathy, exudative diathesis (abnormal permeability of the
capillary walls) and disturbance of the nervous system
(encephalomalacia in chicks). More often sub-clinical vitamin E
deficiency occurs, which is manifest by slow growth, reduced
productivity, frequent health problems and diminished fertility.
Vitamin E in Broilers
Tengerdy et al. (1972) were the first group, reporting a

significantly increased immune response in chicks and hens,
supplemented with elevated dietary levels of vitamin E. Chicks,
infected with E. coli and fed supplemental vitamin E, were found to
have reduced mortality and increased HA titers (Nockels, 1979).
Vitamin E was also demonstrated to induce a higher production of
circulating antibodies against Newcastle disease virus (NDV) and
Pasteurella anatipestifer, following vaccination of broiler chicks
against these antigens (Franchini et al., 1986). In broilers,
immunized against NDV, the highest vitamin E level (300 mg/kg)
in combination with various selenium levels resulted in elevated
body weight gain, favorable feed conversion ratio, significantly

higher antibody titers and improved cellular immune responses
(Swain et al., 2000). In cockerels, fed diets containing either 10 or
300 ppm of vitamin E the ratio heterophils/lymphocyte ratio was
enhanced with higher level suggesting an improved phagocytic
ability of the immune system (Boa-Amponsem et al., 2000). In a
study related to the immune response of broilers to coccidiosis-
immunized chickens, dietary supplementation with Se or vitamin E
reduced mortality and increased body weight gain of non-
immunized chickens infected with E. tenella (Colnago et al., 1984).
In contrast, a later investigation found vitamin E not to counteract
the negative impact of coccidiosis. The likely reason for the
ineffectiveness of vitamin E might be malabsorption during E.
maxima infection, making vitamin E less biologically available to
the infected tissues (Allen and Fetterer, 2002).
Poultry meat has a relatively short shelf life, since oxidation
of lipids give rise to the typical off-odors and off-flavors of spoiled
meat. Vitamin E is nature’s most powerful lipid-soluble antioxidant,
being able to break the free radical induced chain reaction of lipid
oxidation. Numerous studies with meat from poultry, fed on diets
supplemented with elevated levels of vitamin E, have shown that
the oxidative stability of lipids was improved and that the
development of rancid deterioration of the meat was delayed.
Therefore, supplementation of poultry with extra vitamin E is the
most promising approach to achieve and maintain an optimum
quality of poultry meat (Weber, 2001). For this application, the
supplementation levels of vitamin E need to be increased over the
requirement levels for the metabolism, but an important part of this
extra vitamin E is deposited in the meat. Therefore,
supplementation of meat-type poultry with elevated levels of
vitamins improves the nutritional value of the product as well and
can represent a reliable source of essential micronutrients for the
end consumers.
Vitamin E in Breeders and Laying Hens
Supplemental vitamin E was also demonstrated to improve

maternal immunity (Nockels, 1979). Chicks hatched from eggs of
breeders, supplemented with high vitamin E and vaccinated
against NDV showed that vitamin E supplementation of breeder
birds increased the immune response of their progeny (Haq et al.,
1996).
More than 20 years ago, Richter et al. (1986) in absence of
any measurable effects of vitamin E on laying performance, feed
efficiency or mortality, concluded that the natural content (> 1 mg
alpha-tocopherol/kg) of the diet was sufficient for an adequate
performance of laying hens. Later, Scheideler and Froning (1996)
demonstrated that a dietary supplementation of hens with 50
instead of 27 mg/kg improved egg production, which indicated the
essential function of vitamin E for optimizing performance of laying
hens.
In a study, the combination of 65 mg/kg vitamin E and 1’000
ppm vitamin C during heat stress showed the highest lymphocyte
proliferative responses indicating an important function of vitamin
E for the maintenance of health of laying hens (Puthpongsiriporn
et al., 2001). In layers, with chronic heat stress, a vitamin E
supplementation of 500 mg/kg was necessary to alleviate the
adverse effects on egg production and egg weight (Bollengier-Lee
et al., 1998) or 250 mg/kg if the supplemented diet was provided
before, during and after heat stress (Bollengier-Lee et al., 1999).
Vitamin E is a particularly safe micronutrient: hence
supplementation up to 20’000 mg/kg did not impact on
performance of hens and a dose-dependent increase in alpha-
tocopherol concentration in plasma, liver, muscle and egg yolk
was measured (Sünder and Flachowsky, 2001). Higher levels of
vitamin E have been used to improve the oxidative stability of the
egg yolk, particularly when polyunsaturated fatty acids were used.
Grune et al. (2001) added fish oil to the diet of laying hens, which
were supplemented from 0 up to 160 mg vitamin E per kg diet.
Egg yolks contained more n-3 PUFAs, mainly due to accumulation

of docosahexaenoic acid. To prevent the increase of lipid
peroxidation during production and storage of n-3 PUFA-enriched
eggs, a high vitamin E supplementation with at least 80 mg vitamin
E per kg was needed. Only a supplementation of hens with 200
ppm vitamin E reduced significantly lipid oxidation in comparison
to the standard level of 50 mg/kg (Galobart et al., 2001). Eggs are
a perfect vehicle for the transfer of vitamin E: when alpha-
tocopherol - and beta-carotene - were supplemented to laying
hens at dietary levels of up to 400 mg/kg each, their concentration
in chicken egg yolks could significantly be improved. Alpha-
tocopherol increased from the control level of 144 mcg/g of yolk to
477 mcg/g of yolk (at 400 ppm vitamin E) but supplemental beta-
carotene markedly decreased the yolk deposition of alpha-
tocopherol when the two compounds were fed together (Jiang et
al., 1994).
Vitamin K (phylloquinone)
Characteristics and Properties of Vitamin K
Vitamin K1 is present in green plants, green vegetables and

in liver oils. Vitamin K2 has been found in animal and microbial
materials. The main source in animal nutrition is vitamin K3
(menadione). The absorption of vitamin K happens in association
with dietary fats, facilitated by the presence of bile salts. Following
absorption, vitamin K is metabolized in the liver. Very little is
stored; which results in rapid depletion, probably within one week,
if dietary supply is low.
Vitamin K regulates the production of certain coagulation
factors in the blood plasma e.g. prothrombin and clotting factors
VII, IX and X, preventing uncontrolled bleeding from wounds. The
nutritional requirements of poultry are from 2 to 7 mg/kg for the
various poultry categories.
Deficiency of this vitamin increases blood-clotting time, resulting in

hemorrhagic diseases in most tissues and organs. In young
poultry general weakness, rough plumage and paleness as well as
icteric coloration of the comb, the wattles and eyelids because of
anemia have been described. Subcutaneous and intramuscular
hemorrhages and bloody feces due to bleeding in the crop and in
the ceca can occur.
Vitamin K in Broilers
Since vitamin K is an essential factor for blood clotting, the

biological efficacy of various compounds of the vitamin K family
were studied in a bioassay, based on the prothrombin clotting time
of 3-week-old, vitamin-K-depleted male broiler chicks, showing
that menadione (vitamin K3) is bioactive in poultry. (Gropp and
Mehringer, 1990). The interaction between all fat-soluble vitamins
- A, D3, E and K - was studied in broiler chicks, by
supplementation of three levels of each fat-soluble vitamin
representing deficient, optimum and excessive amounts (Abawi
and Sullivan, 1989). Among other findings, the results of this
complex study suggested that higher supplemental levels of
vitamins D and K would improve performance of poultry
occasionally being fed high supplemental levels of vitamins A and
E.
Vitamin K was also found to be important for bone
development. In a broiler study supplemental vitamin K
significantly affected bone quality and feed efficiency. For optimum
bone quality and broiler performance, concentrations of vitamin K
in broilers diets of 8 mg/kg, 2 mg/kg and 2 mg/kg for the starter,
grower and finisher phases, respectively were recommended. The
study showed that the starter period is an important phase for
improving bone quality and validated the mechanism of vitamin K
effects on bone quality, i.e. that vitamin K boosts the carboxylation
of osteocalcin (Zhang et al., 2003).
Vitamin K in Laying Hens and Breeders
Vitamin K is also important in relation to bone formation and

bone re-modeling in poultry egg. Osteocalcin, one of the main
bone proteins - found in bone, in the uterus and in the eggshell - is
dependent on vitamin K and low levels of osteocalcin interfere with
bone mineralization during skeletal development and the eggshell
formation. A link between sub-optimal vitamin K supply and
osteoporosis in laying hens has been proposed by Fleming (2008)
recommending the supplementation of laying feeds with 2 to 3 mg/
per kg diet. Fleming et al. (2003) found that 10 mg/kg resulted in
higher cancellous bone volumes throughout the laying period, also
prolonging the modeling period of bone formation or the inhibition
of medullary bone loss during the first phases of lay. Having an
optimum blood clotting vitamin K supplementation is of practical
importance in birds after beak trimming or in case of nervous
behavior (cannibalism).
Water-Soluble Vitamins
Vitamin B1 (Thiamin)
Characteristics and Properties of Vitamin B1
Thiamin is present in considerable concentrations in yeast,

in cereals, in soya and in oil seeds, but from those feedstuffs, it is
not entirely available to poultry. All vertebrates are almost
completely dependent on dietary vitamin B1 intake, since this
vitamin holds a key function in carbohydrate metabolism and
therefore is present in almost all living cells. Although bacteria are
able to synthesize some thiamin in the intestinal tract (mainly in
the caeca), this material is only accessible to poultry via
coprophagy and accordingly caged laying hens cannot benefit
from that. Commercially vitamin B1 is used as thiamin
hydrochloride or as thiamin mononitrate, both being white
crystalline powders, which in the absence of light and moisture are

fairly stable.
Thiamin is rapidly and actively absorbed from the small
intestine and then transformed by phosphorylation into the active
co-enzyme thiamin pyrophosphate. The body is incapable of
storing free thiamin, but small amounts of the phosphorylated form
is found in all animal cells. Dephosphorylation can occur in the
kidney and excess quantities of the free vitamin are excreted in
the urine.
The active co-enzyme of vitamin B1 is thiamin
pyrophosphate (TPP) involved as a co-enzyme in oxidative
decarboxylation of pyruvic acid and ketoglutaric acid. Poultry of
different categories require a dietary supplementation with vitamin
B1 between 2 to 4 mg/kg feed.
Deficiency symptoms in poultry are loss of appetite, weight
loss, general weakness and death. More particularly, fatty
degeneration and necrosis of heart fibers, bradycardia, heart
failure (Sudden Death Syndrome) have been described and fatty
degeneration and hemorrhages of the liver occur. Mucosal
inflammation, ulcers and hemorrhages of the intestinal tract as
well as skin edema and cyanosis were observed and a
progressive paralysis ultimately leads to death of the birds. In
laying hens, atrophied ovaries were observed, resulting in reduced
egg production.
Vitamin B1 in Broilers
The main investigations on the vitamin B1 requirement of

poultry are quite old and based on this information the
recommendation of NRC (1994) is very low (0.7 mg/kg). However,
a more recent study with broiler chicks indicated that modern
birds, selected for high performance, have a superior requirement
for vitamin B1 (Olkowski and Classen, 1996). Blood thiamin
concentration tended to decline in unsupplemented birds and in
birds supplemented with 8 mg the heart tissue tended to
accumulate thiamin at a considerably higher rate than liver or

brain, indicating that the heart has a higher requirement for thiamin
than other tissues. In a follow-up study (Olkowski and Classen,
1999), it was found that maternal thiamin nutrition substantially
affected thiamin status and thiamin metabolism of the offspring.
The bioavailability of thiamin mononitrate and thiamin
hydrochloride in broiler chickens was found to be equivalent, when
added to a thiamin-deficient diet (Geyer et al., 2000). Withdrawal
of the vitamin premix in the last 21 days of broiler fattening,
combined with heat stress, was found to significantly reduce the
concentration of thiamin (and riboflavin) in the Pectoralis major
muscle (Deyhim et al., 1996).
Vitamin B2 (Riboflavin)
Vitamin B2 is widely distributed in all leafy vegetables, in

meat and in fish. Furthermore, riboflavin is stored in small
quantities in liver, spleen, kidney and cardiac muscle and these
depots are maintained for quite a time. Riboflavin is eliminated in
the urine and the daily losses can amount up to 30% of intake.
The riboflavin requirement in poultry is reported to be between 5
and 12 mg/kg. Laying hens need more vitamin B2 because of the
high production of eggs.
Vitamin B2 occurs as orange-yellow crystals, which are
slightly soluble in water, but insoluble in organic solvents. At
ordinary temperatures, riboflavin is thermostable, remains
unaffected by atmospheric oxygen, but is unstable when exposed
to white or ultraviolet light.
Vitamin B2 is phosphorylated in the intestinal mucosa to
FMN (flavin mononucleotide) during absorption and then
converted to FAD (flavin-adenin dinucleotide) in the liver.
Riboflavin is an essential factor of flavin enzymes (flavoproteins).
These enzymes are involved in the transport and transfer of
hydrogen within the respiration chain and thus contribute to energy

production. Furthermore, they are important for both synthesis and
degradation of fatty acids, which explains why the vitamin B 2
requirement is increased on a high-fat diet. Riboflavin-containing
amino acid oxidases also catalyze the oxidative catabolism of
amino acids. Obvious deficiency signs in poultry are slow growth
and reduced appetite. Mouth, nasal mucous membranes and skin
of the extremities tend to get inflamed and muscular debility,
trembling, spasms and paralysis, due to myelin degeneration of
peripheral nerves can occur. Vomiting, resorption disorders and
diarrhea due to inflammation of mucous membranes of the
digestive tract have been described and in hens egg laying is
disturbed. A reduced hatchability due to high embryo mortality has
been reported as well. The typical manifestation of riboflavin
deficiency in chickens is curled toe paralysis, which are toes bent
inwards.
The severity of vitamin B2 deficiency has been described

with similar clinical signs in two papers. Johnson and Storts (1988)
observed leg weakness and paralysis in chickens on a riboflavin
deficient diet already at 12 days of age. Cai et al. (2009)
demonstrated that dietary riboflavin deficiency causes a
demyelinating peripheral neuropathy in young, rapidly growing
chickens and, for the first time, it was observed that riboflavin
deficiency produces selective injury to peripheral nerve trunks. In
terms of riboflavin requirements an early investigation reported
that under tropical conditions optimum growth of broiler chicks was
attained at a supplementation level of 5.1 mg/kg feed
(Ogunmodede, 1977). Measurement of riboflavin in meat and
organs of the birds showed that increasing levels of dietary
riboflavin increased its deposition in edible tissues of the birds. In
a dose-response study with broiler chicks a severe leg paralysis
and high mortality was observed in those birds, receiving no
supplemental riboflavin (Ruiz and Harms, 1988a). In order to

prevent signs of leg paralysis a dietary level of 4.6 mg riboflavin
per kg of feed was suggested. In confirmation of these earlier
studies, Olkowski and Classen (1998) concluded that the dietary
requirement of riboflavin for fast growing broilers should be set at
a level of 5 mg/kg.
Vitamin B2 in Laying Hens and Breeders
A central physiologic function of riboflavin was described for

the support of the embryonic development (Lee and White, 1996).
Squires and Naber (1993b) investigated the relationship between
dietary riboflavin supplementation of laying hens (1.55-8.8 mg/kg)
and egg production as well as riboflavin content in eggs. Laying
rate, egg weight, hatchability and hen weight were all significantly
depressed by the two lower riboflavin levels later in the
experiment, when compared with the two higher levels. Results
indicated that egg riboflavin concentrations are related to
important production parameters that may be used to predict
future dietary riboflavin inadequacies. A study with breeding hens,
supplemented with 2.5 to 12.5 mg riboflavin per kg and kept under
conditions of a humid tropical environment, confirmed these
findings. Out of the various productive parameters analyzed, the
only one that responded was egg production, increasing
significantly when the dietary riboflavin level was at least 8.5
mg/kg (Arijeniwa et al., 1996).
Vitamin B6 (Pyridoxine)
Vitamin B6 in its main forms pyridoxal and pyridoxamine is

widely distributed in low concentrations in all animals and to a
lower extent cereals and green vegetables. In glycosylated form,
however, it is not entirely bioavailable. Absorption of pyridoxine
occurs in the proximal jejunum. Both forms are bound to albumen

and distributed throughout the animal tissues in form of the co-
enzymes. A large portion of absorbed vitamin B6 might be
excreted in urine. Poultry diets must be supplemented with 3 and 6
mg/kg feed of vitamin B6.
Pyridoxine occurs as colorless crystals, which are soluble in
water and alcohol and resistant to normal heat. Pyridoxal
phosphate and pyridoxamine phosphate are co-factors of various
enzymes – e.g. transaminases, deaminases, desulphydrases and
amino acid decarboxylases – hence are essential for the
metabolism of amino acids, which explains an increased
requirement of vitamin B6 on a high-protein diet. Furthermore,
vitamin B6 is involved in both synthesis and transformation of
glycogen and markedly influences the fatty acid metabolism via
conversion of linoleic to arachidonic acid. Thus, pyridoxine is an
essential element for energy production, fat metabolism, central
nervous system activity and hemoglobin production.
In case of pyridoxine deficiency a loss of appetite, slow
growth, poor feed utilization in poultry has been observed.
Dermatitis, rough and deficient plumage, inflamed edema of
eyelids, anemia and ascites can occur. Through demyelinization of
peripheral nerves, movements look un-coordinated, resulting
ultimately in muscular convulsions, followed by paralysis. In laying
hens impaired egg laying performance, fertility and hatchability will
be induced.
Vitamin B6 plays a central role in many metabolic processes

in poultry, which are responsible for normal growth, the
development of a strong skeleton and a functional nervous system
as well as for disease resistance through an effective immune
response. Accordingly, chicks on a pyridoxine-deficient diet,
developed characteristic vitamin B6 deficiency signs, which were
increased mortality, decreased body weight gain, increased
incidence of abnormal leg conformation and a significant reduction

in antibody levels (Blalock et al., 1984). When broiler chicks were
either fed an adequate or pyridoxine-deficient diet containing
yeast, no weight difference was found between control and B 6-
deficient animals, but the removal of yeast from the diet provoked
a severe symptomatic deficiency (Massé and Weiser, 1994).
Pyridoxine deficiency also influences markedly both ultrastructure
and morphology of the bone (Massé et al., 1994) with markedly
reduced amount of matrix suggesting that there is less collagen
present. It was suggested that pyridoxine is an essential nutrient
for the connective tissue matrix. Since the vitamin B 6-deficient
bones were demonstrated to be osteopenic, it was concluded that
the alterations of the collagen resulted in changes to bone
mechanical performance, although proper cortical bone
mineralization occurred (Massé et al., 1996).
Vitamin B6 in Laying Hens and Breeders
In a classical requirement study with breeding hens the

vitamin B6 needs of the hens was estimated at 1.5 - 2 mg per kg of
feed, based on the reproduction characteristics of the hens, which
included laying performance. A certain benefit of higher pyridoxine
supplementation levels of the hens was observed, when taking
performance characteristics of the chickens into account. (Abend
et al., 1977).
Dietary zinc, along with pyridoxine supplementation (8
mg/kg), increased plasma calcium and phosphorous
concentrations and improved feed efficiency and egg production
(Kucuk et al., 2008). Therefore, this combination was
recommended to improve both performance and egg quality in
laying hens.
Vitamin B12 (Cyanocobalamin)
Vitamin B12 occurs widely in tissues of animal origin and

has important physiological functions. Cyanocobalamine has a
complicated chemical structure and occurs only in animal material
such as liver, kidney and egg yolk and in metabolites of
microorganisms. The microflora of the intestine can synthesize it, if
sufficient cobalt is available. Vitamin B12 is essential for both
growth and health of poultry, but the requirement is rather small,
i.e. between 20 and 40 mcg/ kg of feed. Cyanocobalamine
appears as a dark red crystalline powder, which is slightly soluble
in water and relatively stable to air and heat, but can be destroyed
by light and UV radiation.
Vitamin B12 interacts with the metabolism of folic acid,
participates in the biosynthesis of labile methyl groups and is
necessary for the biosynthesis of purine and pyrimidine bases,
representing essential constituents of nucleic acids. Furthermore,
it is involved in the biosynthesis of methionine and choline.
The most obvious signs of deficiency are impaired
performance, defective feathering, leg weakness, perosis and
gizzard erosion.
Very little research on the beneficial effects of vitamin B 12 in

broiler chicks is reported in the scientific literature. In one study,
vitamin B12 was one component in a complex experimental set-up
with male broiler chicks, which was used to study responses to
dietary supplements of L-methionine, L-cystine, choline and
sulfate as well (Tillman and Pesti, 1986).
Vitamin B12 in Laying Hens and breeders
Vitamin B12 usually does not get a lot of attention, since it is

required only at very low concentrations in the feed. As
microorganism synthesizes cyanocobalamin, coprophagy could
potentially cover a considerable part of the demand of poultry.
However, caged poultry such as laying hens cannot access this
source and thus are entirely dependent on a dietary supply.
Unlike the other B group vitamins, cyanocobalamine can be
accumulated in the tissues, mainly in the liver, but also, to a lesser
degree, in the kidney, muscles, in bone and skin. Nevertheless,
the capacity of the hen to deposit reserves of vitamin B12 is
disputed as well as the question for how long body reserves in
laying hens may support egg laying until they are depleted. An
investigation with graded amounts of dietary vitamin B 12 (4, 8, 16
mcg/kg), given to breeding hens, showed that egg yolk vitamin B12
concentrations were high at the outset, but decreased markedly
within 2 weeks from hens fed the two lowest dietary levels.
Maximum egg production, egg weight, hen weight and hatchability
were obtained when the diet contained 8.0 mcg/kg of vitamin B12.
(Squires and Naber, 1992).
Niacin (Nicotinic acid, Nicotinamide, Vitamin B3)
Characteristics and Properties of Niacin
Nicotinic acid is active in the metabolism as nicotinamide

and represents an indispensable component of hydrogen-carrying
co-enzymes: NAD (nicotinamide adenine dinucleotide) and NADP
(nicotinamide adenine dinucleotide phosphate). Nicotinamide
participates directly in the transfer of hydrogen, which is of utmost
importance in the intermediary metabolism. These biochemical
functions are important for normal tissue integrity, particularly for
the skin, the gastrointestinal tract and the nervous system.
Nicotinamide occurs in all living cells, but is higher in liver

and muscle. An important part of niacin, present in cereals and
their bi-products, is bound and thus not available for absorption by
poultry. The bioavailability of niacin from other feed sources is not
clearly documented. Thus, it is obvious, that a dietary
supplementation of poultry with niacin is necessary. Recent
practical guidelines recommend dietary supplementation levels of
30 to 60 mg niacin/kg feed. Nicotinic acid is a white crystalline
powder, soluble in water and alcohol and stable to atmospheric
oxygen, light and heat.
Clinical deficiency signs in poultry can include loss of
appetite, vomiting, hemorrhagic diarrhea, poor feed utilization and
finally reduced growth. Other symptoms include inflammation of
mucous membranes of the buccal cavity and esophagus, perosis
and deformation of the femur.
Niacin in Broilers
Niacin has been demonstrated in several studies to be

important for growth of broiler chicks. When a diet with a natural
level of 24 mg niacin per kg, which was fortified with 33 mg/kg
niacin according to the recommendations of NRC (1994), was
supplemented additionally with 33 or 66 mg niacin per kg, broiler
chicks responded with improved body weight gains and
ameliorated feed utilization (Waldroup et al., 1985). This result
could be taken as an indication that modern rapidly growing broiler
chicks require higher niacin levels to sustain their high
performance. The two sources of niacin nicotinic acid and
nicotinamide are considered to be equivalent in terms of their
bioefficacy in broiler chicks, fed a maize-soybean meal diet (Ruiz
and Harms, 1988b). Niacin is particularly important for broiler
performance during the starter phase (Ruiz and Harms, 1990;
Celik et al., 2003)
Niacin in Laying Hens
In a study layer performance, feed conversion, hen

nervousness, fertility, and hatchability were recorded, but no
significant differences were found among the dietary treatments
(Ouart et al., 1987). Leeson et al. (1991) evaluated a dietary
supplementation with 22 to 132 ppm niacin in laying hens through
a 364-day trial period. Hens fed 66 or 132 mg/kg supplemental
niacin produced significantly more eggs with less eggshell
deformities than birds fed 22 mg/kg. In a review carried out by
Whitehead (2001) the practical level of supplementation for layers
was reported to be 50 mg/kg feed of niacin. Considering the
progress of breeding since than, the recommendation for modern
laying hens under commercial conditions is most probably higher.
Pantothenic Acid (Vitamin B5)
Characteristics and Properties of Pantothenic Acid
Pantothenic acid occurs only rarely in the free state but it is

widely distributed in all animal and plant tissues as a component of
coenzyme A. Rich sources are yeast, liver, kidney, egg-yolk,
cereals and green plants. Studies with radio-labeled pantothenic
acid in isolated chicken intestine revealed that the absorption in
the digestive tract is occurring through a specific transport
mechanism rather than by simple diffusion (Fenstermacher and
Rose, 1986). The conversion to co-enzyme A (CoA), a carrier
mechanism for carboxylic acids, which represents the conjugated
nucleotide form of pantothenic acid, happens within the tissues.
Acids, when bound to CoA become ‘active’, i.e. have a high
potential for transfer to other groups. The most important form is
acetyl-CoA, which is used in the citric acid cycle when fat,
carbohydrates and certain amino acids are degraded. Acetyl-CoA
is also a precursor for the biosynthesis of long-chain fatty acids,
phosphatides, cholesterol, steroid hormones and bile acids.
Free pantothenic acid is an unstable, extremely hygroscopic

oil. It is therefore unsuitable for practical applications and is used
mainly in the form of calcium and sodium salts. Pantothenic acid is
optically active; only the dextro-rotatory forms, e.g. calcium D-
pantothenate have vitamin activity. The requirements in poultry
range between 10 and 25 mg/kg feed.
Clinical signs of deficiency are reduced growth, decrease in
appetite, poor feed utilization, rough feathering and
depigmentation of the feathers. In severe cases crusts at the
corner of the beak as well as exudate on eye-lids can be seen and
fatty degeneration of the liver occurs. Particularly in layers
impaired laying performance has been described.
Pantothenic Acid in Broilers
Pantothenic acid was demonstrated to influence the energy

utilization in growing chicks (Beagle and Begin, 1976). Germ-free
chicks have a lower requirement for calcium pantothenate (CaPa)
than conventional chicks (Latymer et al., 1981a). When
incorporating copper sulphate in a diet with graded amounts of
calcium pantothenate (CaPa), the Cu supplementation induced
severe signs of pantothenic acid deficiency in those chicks, which
were marginal or deficient in pantothenic acid (Latymer and
Coates, 1981b). It was suggested that high dietary supplements of
CuSO4 induce pantothenic acid deficiency through interference
with the biosynthesis of CoA.
Pantothenic acid in Laying Hens
Requirements of laying hens for this vitamin depend on the

interaction which exists with other vitamins like vitamin C, biotin
and vitamin B12 as well as on the fat content of the ration. Low
levels of vitamin B12 and high levels of fat increase the
requirements of pantothenic acid, while the presence of vitamin C
could reduce dietary needs. But overall, insufficient information is
available to draw final conclusions on the current pantothenic acid

requirement of layers. For an optimum supplementation the
genetic improvements related to performance should be
considered and due to the relationship of pantothenic acid with
energy metabolism, must be adjusted according to the energy
content of the diet.
Folic acid (Pteroylglutamic acid)
Characteristics and Properties of Folic Acid
Folic acid is present in all green-leaved vegetables, also in

liver, kidney, muscle as well as in milk and cheese. Accordingly,
feed ingredients, typically used in poultry nutrition do not provide
adequate folic acid to meet the bird’s requirement. The term
folates embraces a group of biologically active compounds, which
are structurally derived from folic acid. The absorption of folic acid
occurs at the jejunum and thereafter it is transported, attached to a
protein, to the liver. Limited quantities are present in the liver;
otherwise there is no storage of folates in the body. Enzymatic
transformation of folates results into tetrahydrofolic acid (THF),
which is an active co-enzyme. THF is involved in various
biochemical processes, e.g. methylation reactions, amino acid
metabolism as well as biosynthesis of purines and pyrimidines.
The fact that folate activity interacts with many other compounds
of the carbon atom metabolism has to be considered for practical
recommendations of folic acid fortification of poultry diets, which
are generally reported at concentrations of 1,0 to 4.0 mg/kg in the
diet. Folic acid occurs as yellowish-orange crystalline powder,
which is tasteless and odorless. It is slightly soluble in hot water,
but insoluble in alcohol and ether. In crystalline form folic acid is
fairly stable to air and heat, but is degraded by light and ultraviolet
radiation.
Folic acid deficiency result in disturbances of amino acid
metabolism and protein synthesis. Rapidly growing tissues such
as epithelia of the gastrointestinal tract or skin and bone marrow

are particularly affected. Retarded growth, diminished appetite,
rough plumage and feather depigmentation have been described
as well. In severe cases cervical paralysis, leg weakness (perosis)
and white watery diarrhea was observed. In breeding hens,
reduced egg production and hatchability as well as embryonic
malformation may occur.
Folic Acid in Broilers
Wong et al. (1977) set minimum requirement for folic acid

from 0.34 to 0.49 mg folic acid/kg diet for growing broiler chicks. A
close relationship between folic acid requirement and the
availability of choline was observed by Ryu et al. (1995a). Choline
and folic acid both increased tibia length and width and at
adequate dietary levels decreased valgus and varus deformity. A
similar interaction in starting broiler chicks was described between
folic acid and methionine (Ryu et al., 1995b). Folate deficiency
considerably affects various metabolic processes in broiler chicks
and reduced liver dihydrofolate reductase activity (Rennie et al.,
1993) Folic acid has an important function for memory formation in
chicks (Crowe and Ross, 1997).
Folic Acid in Laying Hens
When hens were fed a purified folate-deficient diet (0.07 mg

folate/kg) egg production was found to be reduced. Folates in egg
yolk were concentrated approximately 43-fold relative to the blood
plasma from which they were derived. (Sherwood et al., 1993).
When laying hens were supplemented with 0, 2, 4, 8, 16, 32, 64 or
128 mg/kg of crystalline folic acid for 21 days, laying performance
was not affected but eggs resulted significantly increased in
folates (Hebert et al., 2005). Likewise, in a feeding study with
laying hens 10 mg/kg inclusion of folic acid in the diet resulted in
significantly increased folate incorporation into egg yolk providing
approximately 12.5% of the recommended dietary allowance for

adult humans (House et al., 2002).
The conversion of folic acid (FA) to the biologically active 5-
methyltetrahydrofolate (5-MTHF) is necessary for the deposition of
folate in the egg. When either of these folate forms was used in a
feeding study with laying hens, plasma homocysteine was
decreased, whereas serum and egg folate were increased
(Tactacan et al., 2010).
Biotin (Vitamin H)
Characteristics and Properties of Biotin
Biotin is widely distributed in small concentrations in animal

and plant tissues, but it is poorly bioavailable from cereals, since it
is bound to a protein. Biotin is absorbed in the small intestine,
transported in the blood stream to the various tissues, but almost
not metabolized. Excess biotin is excreted in both urine and feces.
Biotin represents an important co-enzyme in the intermediary
metabolism of carbohydrates, proteins and fats. It is particularly
important for carboxylation reactions, since biotin-containing
carboxylases take up CO2 and transfer it to a suitable substrate. A
possible effect on purine and pyrimidine synthesis has also been
discussed. In its pure form, biotin represents a white crystalline
powder, which is rather stable. The dietary requirement for biotin
in poultry ranges between 150 to 400 mcg/kg of feed.
Biotin deficiency results in retarded growth, reduced
appetite and poor feed conversion. Rough and brittle feathers, dry
skin, dermatitis of foot pads and deformation of the beak can
occur. Fatty liver and kidney syndrome (FLKS) as well as perosis
develop under biotin insufficiency. In breeder hens, poor
hatchability of eggs and malformation of the embryonic skeleton
were observed.
Biotin in Broilers
Although biotin is abundantly present in many feedstuffs,

used in poultry nutrition, this important vitamin is not readily
available from these sources (Blair and Misir, 1989). Biotin
bioavailability was found to be low in most cereal grains, i.e. wheat
(17%), triticale (20%), barley (21%) and sorghum (39%), moderate
in canola meal (66%) and high in soybean meal (98%) and maize
(114%). Accordingly, the biotin requirement of chicks needs to be
covered to a large extent by supplemental biotin. In order to
achieve an adequate biotin status in broiler chicks, plasma and
liver biotin concentrations should be in excess of 4’000 ng/L and
3’000 ng/g, respectively. In classical requirement studies 120 mcg
biotin/kg feed was identified as the minimum dietary requirement
and 160 mcg/kg was required for the prevention of dermatitis,
mortality due to fatty liver and kidney syndrome (FLKS) and leg
deformities (Oloyo and Ogunmodede, 1991; Oloyo and
Ogunmodede, 1992) and 240 mcg biotin/kg were needed since
better results could be obtained (Oloyo, 1994). Biotin was found to
be instrumental for the prevention FLKS (Whitehead et al., 1976).
Biotin deficiency affects the specific activity of the biotin-requiring
enzyme pyruvate carboxylase, which decreased concomitantly
with liver biotin concentration. Addition of supplemental choline to
a biotin-deficient diet was found to decrease the biotin status of
chicks and to increase mortality from FLKS (Whitehead and
Randall, 1982). When dietary biotin was below 200 mcg/kg, the
fatty acid profile in the liver of broilers was significantly altered
(Watkins and Kratzer, 1987). Dietary biotin influences both bone
growth and development as well. Chicks fed biotin-deficient diets
exhibited various deformities, footpad dermatitis, shortened
tibiotarsi and significantly higher bone densities and percentage
bone ash (Bain et al., 1988).
Biotin in Laying and Breeder Hens
Biotin availability from feedstuffs was evaluated in laying

hens, based on determinations of plasma and egg-yolk biotin
(Buenrostro and Kratzer, 1984). Low values were found for wheat
(0%) and sorghum (10 to 20%) and high values for corn (75 to
100%), soybean meal (100%) and meat and bone meal (85%).
Whitehead (1980) found that biotin supplementation during lay did
not have beneficial effects upon egg number, egg size or feed
intake/feed conversion efficiency. However, internal egg quality, as
reflected by albumen height, was slightly improved by biotin. The
concentrations of biotin in the plasma of hens and the hatched
chicks increased with increasing biotin intake of the hen
(Whitehead, 1984). Biotin concentrations in hen plasma and yolk
were linearly related and biotin concentration in chick plasma was
highly correlated with yolk concentration. Production of eggs or
normal chicks was depressed when practical diets were not
supplemented with biotin (Whitehead et al., 1985).
Vitamin C (Ascorbic acid)
Characteristics and Properties of Vitamin C
Vitamin C is widely distributed in high concentrations in

plants, particularly in citrus fruits and green vegetables. Birds are
capable to synthesize ascorbic acid in the kidneys, but in young
chickens and in growing poultry the endogenous production is
insufficient, especially under current intensive husbandry
conditions. Ascorbic acid is readily absorbed in the intestine and
the absorption rate is high (up to 80%). Excess vitamin C or its
metabolites are excreted with the urine.
In its crystalline form, vitamin C is a white to yellow powder,
readily soluble in water but in solution is sensitive to oxygen and
other oxidizing agents. As dietary supplementation 100 to 200
ppm of vitamin C are recommended for the various poultry

categories.
Ascorbic acid is present in all living cells as an important
antioxidant, essential for the formation of intercellular substances
(connective tissue, bones, cartilage) and supports wound healing
via its key function, the hydroxylation of proline to hydroxyproline.
Furthermore, due to its involvement in corticosteroid synthesis it
has an antistress activity and it stimulates defensive mechanisms
such as phagocytic activity of leukocytes and formation of
antibodies.
Clinical vitamin C deficiency does not occur in poultry due
to their ability of endogenous synthesis of ascorbic acid.
Vitamin C in Broilers
Despite poultry have the ability to endogenously produce

ascorbic acid, vitamin C supplementation has been demonstrated
to beneficially influence performance and health of broilers,
particularly under conditions of stress. Interestingly biotin
(Lechowski and Nagórna-Stasiak, 1993) and iron (Nagórna-
Stasiak et al., 1994) were found to influence the vitamin C
synthesis in chickens. Birds fed diets, supplemented with vitamin
C, experienced significant increases in serum levels of ascorbic
acid. It has been reported in several cases, that supplemental
vitamin C improved disease resistance by strengthening the
immune responses (McCorkle et al., 1980; Wu et al., 2000).
Vitamin C in combination with vitamin E reduced mortality from
ascites syndrome as a consequence of an oxidative stress on the
heart of broilers maintained at high altitudes (Villar-Patiño et al.,
2002). Supplemental vitamin C (800 mg/kg) was demonstrated to
positively affect the duodenum and jejunum villus height, width,
surface area and lamina propria thickness, which is an indication
of an improved gut morphology (Zamani Moghaddam et al., 2009).
High levels of vitamin C, supplied to broiler chickens just prior to
slaughter, reduced the stress response of starvation and transport
to the abattoir indicating that ascorbic acid supplementation might

improve the quality of broiler meat. (Satterlee et al., 1989)
Vitamin C in Laying and Breeding Hens
Although, laying hens are not dependent on exogenous

vitamin C, supplemental ascorbic acid demonstrated to have
beneficial effects under stress conditions. In laying hens, kept at
high temperatures and crowding conditions, supplemental vitamin
C (200 ppm) reduced mortality, increased slightly shell weight per
unit surface area and improved the Haugh units (Cheng et al.,
1990). The vitamins C and E, both potent antioxidants, were
demonstrated to act synergistically in heat-stressed laying hens,
when measured by serum levels of tri-iodothyronine (T3) and
thyroxine (T4), which were increased by the supplementation or by
the plasma concentration of adrenocorticotropic hormone (ACTH),
which was decreased (Sahin et al., 2002a). Vitamin C exerted also
beneficial effects under low ambient temperature conditions (Sahin
et al., 2002b). The benefits of vitamin C for the immune response
of laying hens challenged with concanavalin A and Salmonella
typhimurium lipopolysaccharide were also demonstrated in
combination with vitamin E (Puthpongsiriporn et al., 2001). For
these reasons practical recommendations for commercial
husbandry conditions consider also vitamin C for supplementation
of laying hens.
Also in broiler breeders, under hot and humid conditions,
supplemental ascorbic acid has been demonstrated to beneficially
influence egg reproduction in laying hens (Panda et al., 2008).
CONCLUSIONS
It is widely accepted by the poultry industry that the

minimum dietary vitamin levels, required to prevent clinical
deficiencies, does not support optimum health, performance and
welfare of poultry. The reasons for that are manifold: (1) genetic
improvement of the breeds improve continuously and modern

genetics achieve higher performance with less feed intake; (2)
modifications in nutrition, management and husbandry, tend to
increase the demand for vitamins per unit output; (3) the intensive
poultry production may generate a certain level of metabolic,
social, environmental and disease stresses, causing sub-optimal
performance and higher susceptibility to vitamin deficiencies; (4)
the contamination of the feed with mycotoxins and vitamin
antagonists can limit or even block the action of certain vitamins.
Any of those factors can separately or collectively affect the need
for each vitamin. As level, intake and utilization of vitamins from
natural sources is unpredictable, it is safer to cover the entire
vitamin requirement of poultry through dietary supplementation.
Since recent research on the vitamin requirements of modern
birds is limited, breeding companies and vitamin manufacturers
regularly publish guidelines, which consider all important
developments in vitamin knowledge in order to support poultry
nutritionists in their task to formulate the most adequate diets for
all poultry categories. These recommendations for vitamin
supplementation generally exceed the minimum requirements,
postulated by governmental institutions, since they aim at entirely
exploiting the productivity potential of modern birds under today’s
husbandry conditions.
Optimum Vitamin Nutrition (OVN) is the recommended
vitamin supplementation suggested by DSM Nutritional Products
since 1958. The suggested levels and ratios amongst vitamins,
based on both research data, genetic companies and industry
experience, are designed to optimize birds’ performance, health,
welfare and quality of animal origin foods. For certain vitamins
supplementation at higher levels is suggested for achieving
additional effects, such as modulation of the immune response
and subsequent strengthening of the disease resistance or the
improvement of meat quality. Finally, dietary vitamins are
deposited in meat and eggs, which increase the nutritional value of
these important food products. All these factors need to be taken
into account, when recommendations for the correct

supplementation levels of vitamins are given.
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NUTRITIONAL REQUIREMENTS OF AMINO ACIDS
FOR GROWING PIGS
John K. Htoo
Evonik Nutrition & Care GmbH
Rodenbacher Chaussee 4, 63457 Hanau, Germany
ABSTRACT
In response to continuing improvement in the lean gain

potential of modern pig genetics coupled with increased
application of low protein-amino acids fortified diets and a
decrease use of in-feed antimicrobial growth promoters (AGP),
numerous studies have been conducted to determine the
requirements of amino acids for growing pigs during the last
decade.
This review covers more than 75 papers published during
the last 15 years and provides an update on the requirements of
amino acids for growing pigs with body weight of approximately 25
to 115 kg. Lysine requirement has increased over the years for
today’s high lean pigs which averaged 20 g of SID lysine/kg of
gain for 25-50 kg pigs and 21g of SID lysine/kg of gain for 50-115
kg pigs. The optimal ratios of amino acids relative to lysine are
slightly higher than previously assumed values which may be
attributed to differences in health status associated with decreased
use of AGP in the diets. These data could be used for balancing
according to the “Ideal Protein” concept thereby optimizing the
utilization of amino acids, and reducing excretion of excess protein
to the environment.
INTRODUCTION
To achieve the best possible production efficiency, one of

the current aims of pork industry is to maximize lean gain of pigs
by using high lean genetics and through proper nutrition. As such,
today’s modern pigs are leaner and grow faster compared with
pigs of about a decade ago. Lean gain can be maximized only

when adequate quantities of nutrients, particularly essential amino
acids (AA) and energy, are supplied in the diet. For doing so,
swine nutritionists need to know the requirements of AA which can
be influenced by many factors, including dietary crude protein (CP)
level, energy density, genetics, sex, health status and
environmental conditions.
Lysine (Lys) is the first limiting AA in typical swine diets.
With increases in lean gain capacity of the pigs, the requirement of
Lys will also increase because the primary role of Lys is for body
protein (lean gain) deposition. In commercial diet formulations, it is
a common practice to balance the dietary level of other essential
amino acids (EAA) relative Lys on the basis of an ideal protein (IP)
ratio. The first proposal for IP concept referred directly to the ratio
of essential AA in the diet without any access or deficiency. Fuller
et al. (1989), and Chung and Baker (1992) further developed the
IP concept wherein the concentrations for each of other EAA are
expressed as a percentage of Lys, which is set at 100%. The
advantage of IP concept is that the requirements of other EAA can
be estimated if Lys requirement is known. Valine (Val) becomes
the next limiting AA after Lys, threonine (Thr), methionine (Met)
and tryptophan (Trp) in low CP diets for pigs. After Val, isoleucine
(Ile) becoming limiting or in some finisher pig diets, they are co-
limiting. For accurate balancing of dietary AA level, it is crucial to
have knowledge about the optimal IP ratios for pig diets.
Pigs are often exposed to chronic subclinical level of
diseases and environmental stress in commercial farms, resulting
in a reduced feed intake and impaired performance. Such immune
challenges have increased after the ban of antimicrobial growth
promoters (AGP) in animal feeds in the European Union in 2006.
During immune challenges, nutrients are redirected away from
growth and towards tissues involved in immune response. This
implies that the requirement for some AA for the production of
compounds that are involved in the immune response such as
acute phase proteins will be increased. Thus, it is possible that IP
ratios for some AA relative to Lys may be increased when pigs are
exposed to immune challenge conditions and when AGP is not
included in the diets.
Different approaches could be used for estimating the
requirement or optimal ratio of AA. The most commonly used
method is the empirical dose-response method wherein AA
requirement is estimated based on the optimal growth responses
to graded levels of the test AA (1st limiting). Another approach is
based on the factorial approach which combines the AA
requirements for maintenance and protein deposition for a given
body weight (BW) of pigs (NRC, 2012). Other methods used to
estimate AA requirements include the deletion method (Wang and
Fuller, 1989) and using the response of plasma urea nitrogen
concentration as indirect indication of dietary AA supply (Zhang et
al., 2012).
The aim of this article was to review and provide an update
on the requirements of AA for growing pigs with BW approximately
25 to 115 kg based on the published empirical data during the last
15 years. For Lys, the requirements per se expressed in
standardized ileal digestible (SID) % of diet and in SID g/kg gain
will be reported. However, for the other EAA, i.e. Thr, SAA, Trp,
Val, Ile and Leu, the requirements will be expressed as optimal
ratio (%) relative to dietary SID Lys concentrations. Due to the
limited space, the estimation of economic optimum ratios of AA is
not included in this review.
LYSINE REQUIREMENTS FOR GROWING PIGS
Lysine is the first limiting AA in typical diets for pigs. The

primary role of Lys is for body protein synthesis. A portion of the
dietary Lys is also utilized for obligatory oxidation, which occurs
even when Lys intake limits protein deposition, is a component of
the Lys requirement in animals. The estimates of obligatory Lys
oxidation in growing pigs range from 15 to 30% of available Lys
intake (Moehn et al., 2004). A considerable amount of research
has been done to re-evaluate Lys requirements of growing and

finishing pigs in the last decade. In the experiments included in
this review, Lys was the AA under investigation and all other AA
met or exceeded the requirements.
Growing pigs (25 to 50 kg BW)

The summary of Lys requirement estimates for 25-50 kg
growing pigs as well as the experimental conditions and the
regression models applied is given in Table 1.
Table 1 - Review of lysine requirements for 25-50 kg growing pigs

BW Sex1 Optimal Optimal SID Model3 Response Reference
Breed2
performance3 Lys
kg ADG ADFI G:F % diet mg/g criteria
ADG
15-30 B HLG 771 1066 0.72 1.10 15.2 Q ADG Oliveira et al. 2006
24-62 B HLG 956 2065 0.46 0.90 19.4 QBL ADG Fortes et al., 2011
25-50 M HLG n/a n/a n/a 1.04 n/a BL, Q ADG, G:F Landero et al., 2016
25-59 G HLG 850 1758 0.48 1.05 21.7 QBL ADG Gattas et al., 2012
25-65 B HLG 1000 2156 0.46 0.90 19.4 anova ADG, G:F Haese et al., 2011
30-60 B HLG 1000 1900 0.53 1.02 19.38 BL ADG, G:F Oliveira et al. 2009
30-60 B HLG 1023 1967 0.52 1.05 20.19 BL ADG, G:F Abreu et al. 2007a
31-50 M PxS 621 1330 0.47 1.07 22.9 Q ADG, G:F Warnants et al. 2003
25-55 1.02 19.8 Average
0.98 NRC, 2012
1
B = barrows, G = gilts, M = mixed-sex.
2
HLG = High lean genetics; D = Duroc; P = Pietrain; S = Seghers.
3
n/a = Not available.
4
Q = quadratic; QBL = quadratic broken-line; BL = linear broken-line, anova = Analysis of
variance.
Oliveira et al. (2006) determined the standardized ileal

digestible (SID) Lys requirement for 15-30 kg high lean barrows
using corn-soybean meal (SBM)-based diets. Based on the
optimum average daily gain (ADG) responses, the SID Lys
requirement was 1.10% for these pigs. Fortes et al. (2011) and
Haese et al. (2011) estimated the SID Lys requirement to optimize
the ADG of approximately 25-65 kg pigs to be 0.90%. More
recently, Landero et al. (2016) reported that the SID Lys
requirement was 1.04% to optimize the ADG and G:F of 25-50 kg
pigs. To optimize ADG, Gattas et al. (2012) estimated a SID Lys

requirement of 1.05% for 25-59 kg high lean pigs. Based on ADG
and G:F responses, the SID Lys requirement of 30-60 kg high lean
pigs was estimated at 1.02 and 1.05% (Oliveira et al. 2009; Abreu
et al. 2007a). Warnants et al. (2003) reported that the SID Lys
requirement was still higher than 1.07% to optimize the ADG and
G:F of 31-50 kg Pietrain pigs.
Based on these results, the average SID Lys requirement is
1.02%, which corresponds to 19.8 mg SID Lys/g of ADG for 25-50
kg high lean pigs. This estimate is rather close to the NRC (2012)
recommendation of 0.98% SID Lys.
Growing pigs (50 to 75 kg BW)
The Lys requirement estimates for 50-75 kg growing pigs

are summarized in Table 2.
Table 2 - Review of lysine requirements for 50-75 kg growing pigs

BW Sex1 Optimal Optimal SID Model3 Response Reference
Breed2
performance3 Lys
kg ADG ADFI G:F % diet mg/g criteria
ADG
35-60 G PIC 930 1944 0.48 1.02 21.7 anova ADG Main et al. 2008
43-70 B PIC 992 2111 0.47 0.91 19.4 anova ADG, G:F Main et al. 2008
38-65 G PIC 950 1920 0.49 1.04 21.8 BL ADG, G:F Shelton et al. 2011
40-70 B DxLR/Y 879 2020 0.44 0.91 20.9 anova ADG, G:F Deng et al. 2010
41-69 B PxS 866 2225 0.39 0.89 22.9 Q ADG, G:F Warnants et al. 2008
41-69 G PxS 789 1850 0.43 1.07 25.1 Q ADG, G:F Warnants et al. 2008
45-70 M LR/LWxP 815 1827 0.45 0.89 20.0 anova ADG, G:F Nery et al., 2012
50-75 M PIC n/a n/a n/a 0.84 n/a QBL, BL ADG, G:F Landero et al., 2016
55-80 G PIC 970 2110 0.46 0.90 19.6 BL ADG Shelton et al. 2011
60-97 B HLG 1198 2828 0.42 0.90 21.2 Q ADG, G:F Abreu et al. 2007b
60-85 G PIC 974 2322 0.42 0.88 21.0 anova ADG, G:F Main et al. 2008
69-93 B PIC 946 2350 0.40 0.88 21.9 anova ADG, G:F Main et al. 2008
60-96 B PIC 1136 2847 0.40 0.85 21.3 Q G:F Oliveira et al., 2014
50-75 0.92 21.4 Average
0.85 NRC, 2012
1
2
HLG = High lean genetics; D = Duroc; P = Pietrain; S = Seghers, Y = Yorkshire.
3
4
variance.
Main et al. (2008) determined the SID Lys requirement of

approximately 35-70 kg PIC pigs fed corn-SBM-based diets. The
best ADG and G:F were observed with the dietary SID Lys level of
1.02% for gilts and 0.91% for barrows, respectively. Similarly,
Shelton et al. (2011) estimated the SID Lys requirement for 38-65
kg PIC gilts to be 1.04% to optimize the ADG and G:F in a
commercial farm. Deng et al. (2010) also estimated the SID Lys
requirement for 40-70 kg crossbred barrows fed corn-SBM-based
diets to be 0.91% to optimize the ADG and G:F responses.
Warnants et al. (2008) determined the SID Lys requirement of 41-
69 kg high lean Pietrain pigs. The best ADG and G:F were
observed with a dietary SID Lys level of 0.89% for barrows and
1.07% for gilts, respectively. Based on ADG and G:F, Nery et al.
(2012) reported the optimal dietary SID Lys to be 0.89% for 45-70
kg Pietrain pigs.
More recently, Landero et al. (2016) reported that the SID
Lys requirement was 0.84% to optimize the ADG and G:F of 50-75
kg PIC pigs. Shelton et al. (2011) estimated the SID Lys
requirement of 55-80 kg PIC gilts fed corn-SBM diets to be 0.90%
to optimize the ADG and G:F in a commercial farm. Similarly,
Abreu et al. (2007b) estimated the SID Lys requirement for 60-97
kg high lean barrows fed corn-SBM-based diets to be 0.90% to
optimize the ADG and G:F responses. Main et al. (2008)
determined the SID Lys requirement of high lean PIC growing pigs
with BW ranged from 60 to 93 kg. The best ADG and G:F were
observed with the dietary SID Lys level of 0.88% for both barrows
and gilts. To optimize the G:F of 60-96 kg PIC barrows, Oliveira et
al. (2014) estimated the SID Lys requirement to be 0.85%.
Overall, the average SID Lys requirement estimate is
0.92%, which corresponds to 21.2 mg SID Lys/g ADG for 50-75 kg
high lean pigs. This estimate is slightly higher than the NRC
(2012) recommendation of 0.85% SID Lys.
Finishing pigs (80 to 115 kg BW)
The Lys requirement estimates for 80-115 kg finishing pigs

are summarized in Table 3. Cline et al. (2000) assessed the
dietary Lys requirement of finishing gilts with BW ranged from 55
to 116 kg at 8 different research locations. Based on the ADG and
G:F responses, the SID Lys requirement for these gilts was
estimated at 0.73%. Main et al. (2008) also determined the SID Ile
requirement of 78-103 kg PIC barrows fed corn-SBM-based diets.
The ADG and G:F were optimized at the dietary SID Lys level of
0.72% for these pigs. Similarly, Shelton et al. (2011) estimated the
SID Lys requirement of 84-110 kg PIC gilts fed corn-SBM diets to
be 0.89% to optimize the ADG and G:F in a commercial farm. In
95-122 kg PIC barrows fed corn-SBM-based diets, Arouca et al.
(2007) estimated the SID Lys requirement to be 0.63% to optimize
the ADG and G:F. Santos et al. (2011) estimated the SID Lys
requirement for 95-125 kg PIC barrows fed corn-SBM diets to be
0.74% to optimize the ADG and G:F responses.
Table 3 - Review of lysine requirements for 80-115 kg finishing pigs

Optimal Optimal SID
BW Sex1 Breed2 Model3 Response Reference
performance Lys
kg ADG ADFI G:F % dietmg/g ADG criteria
55-116 G n/a 850 2730 0.31 0.73* 23.5 QBL ADG Cline et al. 2000
78-103 G PIC 916 2545 0.36 0.72 20.0 anova ADG, G:F Main et al. 2008
84-110 G PIC 980 2540 0.39 0.89 23.0 BL ADG Shelton et al. 2011
*
95-122 B PIC 1095 3508 0.31 0.63 20.3 Q ADG, G:F Arouca et al. 2007
95-125 B PIC 1081 2940 0.37 0.74 20.6 QBL ADG Santos et al. 2011a
80-115 0.74 21.5 Average
0.67 NRC, 2012
1
2
3
Q = quadratic; QBL = quadratic broken-line; BL = linear broken-line, anova = Analysis of variance.
*
The SID Lys was calculated from total Lys content assuming 88% digestibility.
Based on these results, the average SID Lys requirement

estimate is 0.74%, which corresponds to 21.5 mg SID Lys/g ADG
for 80-115 kg high lean pigs. This estimate is slightly higher than
the NRC (2012) recommendation of 0.67% SID Lys for pigs of

similar BW range.
Optimal SID:NE ratios for growing pigs
Not only dietary AA but energy concentration also plays an

important role in the depositions of body fat and lean tissue.
Research showed that inadequate or excessive of Lys to energy
ratio negatively affect pig performance (Campbell and Taverner,
1988). Thus, the optimal dietary Lys level is also commonly
expressed as relative to the dietary energy content. Because
energy is the single most expensive portion of the feed cost, it is
important to evaluate the optimal level of balanced AA and energy
density in the diet to maximize lean tissue growth and carcass
quality of the pigs.
A considerable amount of research has been done looking
at the effects of Lys to energy ratio on performance and carcass
quality. However, most of these studies were based on
metabolizable energy (ME) system, which does not take the
energy loss as heat into account. The net energy (NE) system on
the other hand considers the amount of heat lost during digestion
and subsequent deposition of nutrients in protein and fat tissue.
Thus, NE provides more accurate estimates of the energy in an
ingredient or diet that is actually available to the pig. However,
limited information is available on the NE requirements and
optimum SID Lys:NE ratio of growing-finishing barrows fed low CP
diets supplemented with AA.
Zhang et al. (2011) investigated the effects of NE contents
and the SID Lys:NE ratios on the performance and the carcass
characteristics of growing-finishing pigs fed low CP diets in a 3 × 3
factorial arrangement having 3 NE densities and 3 SID Lys:NE
ratios for both growing (23-56 kg BW) and early finishing phase
(62-99 kg BW). The CP content of the diets was approximately
14% in the grower diets and 11% in the finisher diets.
Table 4 - Effects of NE content and SID Lys:NE ratios on pig

performance (Zhang et al. (2011)
Growing phase (23-56 kg BW)
NE level (MJ/kg) SID Lys:NE (g/MJ) P-value
9.66 9.87 10.08 0.89 0.96 1.03 NE Lys:NE
ADG, g 709b 740a 720ab 712b 714b 743a 0.04 0.03
ADFI, g 1540 1543 1510 1523 1523 1545 ns ns
FCR, g/g 2.17a 2.09b 2.10b 2.14a 2.13a 2.08b <0.01 <0.01
Finishing phase (62-99 kg BW)
NE level (MJ/kg) SID Lys:NE (g/MJ) P-value
9.83 10.04 10.25 0.69 0.76 0.83 NE Lys:NE
b a a b a a
ADG, g 853 903 918 856 915 903 0.03 0.02
ADFI, g 2471 2433 2512 2397 2541 2477 ns ns
FCR, g/g 2.90a 2.69b 2.73b 2.81 2.77 2.75 <0.01 ns
a,b
Means in a row with different letters are different (P < 0.05); ns = non-significant.
During the growing phase (23-56 kg BW), the best ADG

and FCR were observed for the barrows that received the diet
containing 9.87 MJ/kg. No further improvements were found for
10.08 MJ/kg NE (Table 3). There was a linear effect of SID Lys:NE
ratio on ADG and FCR which were optimized at the highest SID
Lys:NE (1.03 g/MJ). Based on the optimal NE (9.87 MJ/kg or 2.36
Mcal/kg) and SID Lys:NE ratio (1.03 g/MJ), the optimal dietary SID
Lys level is estimated to be 1.02%, which is in good agreement
with the NRC (2012) recommendation of 0.98%. For the finishing
phase (62-99 kg BW), best performance was achieved with the
dietary NE of 10.04 MJ/kg (2.40 Mcal/kg). The effect of Lys:NE
was not significant but seemed to optimize at the dietary SID
Lys:NE of 0.76 g/MJ, indicating that the SID Lys requirement for
these pigs (62-99 kg BW) is at least 0.76% which is close to the
NRC (2012) value of 0.73%.
OPTIMAL THREONINE TO LYSINE RATIOS FOR

GROWING PIGS
Threonine (Thr) is usually the second- or third-limiting AA in

typical cereal-based swine diets. In addition to its primary role in
protein synthesis, Thr is involved in physiological functions of the
gut. Among the essential AA, Thr is particularly important for

synthesis of intestinal mucosal proteins and mucins and thus,
plays an important role in the maintenance of intestinal health and
gut barrier integrity (Bertolo et al. 1998).
The optimal SID Thr:Lys ratio estimates for 50-75 kg
growing-finishing pigs are summarized in Table 5.
Table 5 - Review of optimal SID Thr:Lys ratios for growing-finishing

pigs
BW Sex1 Breed2 Lys Thr:Lys NDF3 AB4 Optimal Model5 Response Reference
kg (SID, %) % Thr:Lys criteria
SID, %
22-50 M DxLW/LR 0.90 55 to 75 n/a (-) 70 BL, Q ADG, G:F Zhang et al., 2013
25-50 G Gen 0.9 45 to 90 7.8 (-) 65 BL, Q ADG, G:F Mathai et al, 2016
25-50 G Gen 0.9 46 to 90 16.7 (-) 67 BL, Q ADG, G:F Mathai et al, 2017
Buraczewska et al,
25-52 B HLG 0.87 53 to 70 n/a (-) 65 anova ADG, NR6
2006
30-50 M LR/LWxP 0.80 42 to 62 n/a (-) 63 BLQ ADG, G:F Kluge et al., 2002
35-65 M LRxP 0.77 54 to 71 n/a (-) 69 BL ADG Ettle et al, 2004
25-50 66 Average
60 NRC, 2012
65-100 M LR x P 0.61 54 to 72 n/a (-) 69 QBL ADG Ettle et al, 2004
72-104 B LW/LR 0.61 56 to 77 n/a (-) 71 BL, Q ADG, G:F Xie et al., 2014
90-118 G DxLW/LR 0.51 54 to 78 n/a (-) 67 BL, Q ADG, G:F Ma et al, 2015a
95-125 B PIC 0.81 58 to 73 n/a (+) 65 Q G:F Santos et al., 2010
80-115 68 Average
63 NRC, 2012
1
2
D = Duroc; LW = Large white, LR = Landrace, HLG = High lean genetics; P = Pietrain;
S = Seghers, Y = Yorkshire, Gen = Genetiporc.
3
4
AB = Antibiotics: (-) refers to without AB and (+) means AB was added in the diet.
5
variance.
6
NR = Nitrogen retention.
Zhang et al. (2013) observed a clear response of ADG and

G:F by increasing SID Thr:Lys ratio from 55 to 75% in low CP-
AGP free diets fed to 25-50 kg pigs raised under commercial
conditions. The average SID Thr:Lys ratio to optimize ADG and
G:F was 70% based on broken-line and quadratic regression.
More recently, Mathai et al. (2016) reported that the average
optimum SID Thr:Lys ratio for ADG and G:F of 25-50 kg pigs was
67% when fed high fiber diet (contained 16.7% NDF) while 65%
was optimal for pigs fed a low fiber diet (7.8% NDF) indicating that
feeding diets high in fiber increases the demand for Thr for mucin
production relative to body growth in pigs. Buraczewska et al
(2006) also reported that the ADG and N-retention (g/d) of 25-52
kg pigs maximized at 65% SID Thr:Lys. Based on the ADG and
G:F responses, Kluge et al. (2002) estimated the optimal SID
Thr:Lys ratio for 30-50 kg crossbred pigs to be 63%.
Based on ADG response, Ettle et al (2004) estimated a SID
Thr:Lys ratio of 69% for 35-65 kg pigs and 69% for 65-100 kg. Xie
et al. (2014) estimated the SID Thr:Lys ratio to optimize ADG and
G:F of 65-100 kg pigs to be 69%. For 90-115 crossbred barrows,
Ma et al. (2015a) reported that ADG and G:F were optimized at a
SID Thr:Lys ratio of 71%. Santos et al. (2010) estimated the SID
Thr:Lys for 95-125 kg PIC barrows fed corn-SBM diets to be 65%
to optimize the G:F.
Based on these results, the growth performance of high
lean pigs of 25-50 kg and 80-115 kg maximized at an average SID
Thr:Lys ratio of 66 and 68%, respectively. These estimates are
slightly higher than the NRC (2012) recommendations of 60 and
63% for the corresponding BW ranges.
Use of dietary AGP helps maintaining animal’s health
status. Feeding animals AGP-free diets may allow higher microbial
growth in the gut and potentially affect the gut health and AA
utilization. Bikker et al., (2007) reported that the SID Thr:Lys ratio
to optimize ADG was higher (71%) for 25-110 kg pigs fed AGP-
free diet compared with those fed AGP-added diet (65%; Figure
1). It is possible that more dietary Thr is utilized to enhance the
immune system through its incorporation into immunoglobulin (Li
et al., 1999).
Figure 1 - Effect of dietary SID Thr:Lys ratio on average daily

(ADG) gain of pigs fed diets with or without AGP (Bikker
et al., 2007).
OPTIMAL SULFUR-CONTAINING AMINO ACIDS (SAA) TO

LYSINE RATIOS FOR GROWING PIGS
Methionine (Met) is considered to be the 2nd or 3rd limiting

AA in typical swine diets. Methionine is an essential AA for normal
growth which cannot be synthesized in the body, whereas cysteine
(Cys) can be converted from Met as need. Hence, the amount of
Met needed in the diet depends on the amount of Cys present.
Thus, it is also important to have requirement estimates for
Met+Cys or sulfur amino acid (SAA) in addition to Met when
formulating swine diets.
Only a few studies have investigated the optimum SID
SAA:Lys ratios in diets for growing-finishing pigs (Table 6). Zhang
et al (2015) reported that the ratio of SID SAA:Lys required to
optimize ADG and FCR of 25-50 kg pigs fed low CP diets and
raised under commercial conditions was 63% based on broken-
line and quadratic broken-line regressions. Gaines et al. (2004)

observed an increased in ADG and G:F by increasing SID
SAA:Lys ratio from 50 to 70% in 29-45 kg pigs. Based on broken-
line regression the average optimal SID SAA:Lys ratio was
estimated to be 61%. Similarly, Yi et al. (2005), based on broken-
line and quadratic regression analysis of ADG and G:F responses,
estimated the average optimal SID SAA:Lys ratio of 61% for both
31-49 kg PIC gilts and barrows.
For 53-105 kg Pietrain pigs, the quadratic regression
estimated an average optimal SID SAA:Lys ratio of 61% for ADG
and FCR (Roth et al., 2000). Pena et al. (2008) estimated the SID
SAA:Lys ratio for 84-110 kg PIC barrows fed corn-sorghum-
soybean meal based diets supplemented with ractopamine and
AGP was 54% to optimize ADG but a higher ratio of 66% was
needed to maximize cholesterol concentration in loin muscle,
which provide an average ratio estimate of 60%. Santos et al.
(2011b) also fed corn-sorghum-soybean meal diets supplemented
with AGP to 95-125 kg PIC barrows and observed that the ADG
and FCR were optimized at a dietary SID SAA:Lys of 57%. More
recently, Ma et al. (2016) estimated the SID SAA:Lys ratio to
optimize ADG and FCR of 96-120 kg gilts to be 60%.
Based on these literature data, the average SID SAA:Lys
ratio to optimize the growth performance was estimated at 61% for
25-50 kg pigs and 60% for 80-115 kg pigs. These estimates are
slightly higher than the NRC (2012) recommendation of 56 and
58% for the corresponding BW range.
The differences of SAA:Lys ratio may be attributed to
differences in health status (with or with AGP) and genetics (lean
gain potential) of the pigs used and the regression methods
applied in these studies. Dietary Met can be converted to supply
Cys which is the rate-limiting substrate for the synthesis of
glutathione (GSH) which is the major intracellular antioxidant
involved in immune functions. In a N-balance study with growing
pigs, Litvak et al. (2013) showed that immune challenge by
lipopolysaccharide (LPS) injection reduced body protein deposition
rate but the optimal dietary Met to Met+Cys ratio for maximum
body protein deposition increased from 57 to 59%. Similarly, the
dietary SID SAA:Lys to maximize body protein deposition
increases from 55 to 75% when growing pigs are immune
challenged with LPS (Kim et al., 2012). These results indicate that
the needs of Met+Cys, including Met requirement for converting to
Cys, are increased during immune challenge.
Table 6 - Review of optimal SID SAA:Lys ratios for growing-

finishing pigs
BW Sex1 Breed2 Lys SAA:Lys AB3 Optimal Model4 Response Reference
kg (SID, %) SAA:Lys criteria
SID, %
20-50 M DxLW/LR 0.90 50 to 70 (-) 63 BL, QBL ADG, FCR Zhang et al., 2015
29-45 M PIC 0.90 50 to 70 (-) 61 BL ADG, FCR Gaines et al., 2004
31-49 G PIC 0.90 50 to 70 (-) 61 BL, Q ADG, G:F Yi et al., 2005
31-49 B PIC 0.90 50 to 70 (-) 61 BL, Q ADG, G:F Yi et al., 2005
25-50 61 Average
56 NRC, 2012
53-105 M LR x P 0.48 44 to 69 (-) 61 Q ADG, FCR Roth et al., 2000
84-110 B PIC 0.94 54 to 66 (+) 60 anova ADG, CH5 Pena et al., 2008
95-125 B PIC 0.75 57 to 73 (+) 57 anova ADG, FCR Santos et al. 2011b
96-120 G DxLW/LR 0.51 48 to 68 (-) 60 BL, Q ADG, FCR Ma et al., 2016
80-115 60 Average
58 NRC, 2012
1
2
D = Duroc; LW = Large white, LR = Landrace.
3
4
variance.
5
CH = cholesterol concentration in loin muscle.
OPTIMAL TRYPTOPHAN TO LYSINE RATIOS FOR

GROWING PIGS
Tryptophan (Trp) is required not only for protein synthesis

but also involved in various metabolic pathways including immune
response and the formation of serotonin which involves in feed
intake regulation (Henry et al. 1992). Tryptophan is usually
considered the 4th limiting AA in typical cereal-based swine diets.
A moderate amount of research has been conducted to evaluate
the Trp:Lys ratios for growing-finishing pigs in the last decades.

The summary of optimal SID Thr:Lys ratio estimates for growing-
finishing pigs are summarized in Table 7.
Table 7 - Review of optimal SID Trp:Lys ratios for growing-

finishing pigs
BW Sex1 Breed2 Lys Trp:Lys AB3 Optimal Model4 Response Reference
kg (SID, %) Trp:Lys criteria
SID, %
20-55 M HLG 0.82
16.9 to 22.2 (-) 21 BL ADG, FCR Millet et al. 2015
25-40 M LW/LRxD 0.66
12.8 to 17.9 (+) 18 BL, QBL ADG, G:F Quant et al, 2012
25-40 M LW/LRxD 0.66
13.1 to 18.1 (+) 17 BL, Q ADG Quant et al, 2012
25-50 G LR/LWxP 0.87
12.8 to 23.1 (-) 21 Expo ADG, FCR, NR Eder et al. 2003
25-50 M D x LW/LR 0.90
13 to 25 (-) 21 BL, BLQ ADG, FCR Zhang et al. 2012
Van der Aar et al.
25-55 G Yx LR 0.87
15 to 24 (-)
20 BL ADG, FCR, FI 2012
25-45 20 Average
17 NRC, 2012
Van der Aar et al.
55-110 G Yx LR 0.55 15 to 24 (-) 19 BL ADG, FCR, FI
2012
66-124 M PIC 0.78
15 to 19.5 (-) 19.5 anova ADG, FCR Salyer et al. 2013
67-96 B LWxLR 0.61
13.1 to 26.2 (-) 22 BL, Q ADG, FCR Xie et al. 2014
80-105 B LW/LRxD 0.52
11.5 to 23.1 (-) 19 BL ADG, FCR Guzik et al. 2005
5
80-115 G LR/LWxP 0.56
12.3 to 22.9 (-) 20 Expo. ADG, FCR, NR Eder et al. 2003
89-114 B NH 0.55
10.9 to 21.8 (-) 15 anova ADG, FCR Kendall et al. 2007
91-123 B PIC 0.55
13.0 to 23.5 (+) 20 anova ADG, FCR Kendall et al. 2007
99-123 B PIC 0.55
13.0 to 21.0 (-) 17 anova ADG, FCR Kendall et al. 2007
89-121 G DxLW/LR 0.51
12 to 24 (-) 18 BL, Q ADG, FCR Ma et al. 2015b
Goncalves et al.
107-125 G PIC 0.75
14.5 to 24.5 (-)
19.8 BLQ ADG, FCR 2015
80-115 19 Average
18 NRC, 2012
1
2
HLG = High lean genetics, D = Duroc; LW = Large white, LR = Landrace, Y =
Yorkshire, NH = Newsham genetics.
3
4
Q = quadratic; QBL = quadratic broken-line; BL = linear broken-line, Expo =
exponential, anova = Analysis of variance.
5
NR = Nitrogen retention.
Millet et al. (2015) reported the average SID Trp:Lys ratio to

optimize the ADG and FCR of 20-55 kg high lean gain pigs based
on broken-line regression was 21%. Quant et al. (2012) evaluated
the effect of ingredient composition on Trp:Lys ratio of 25-40 kg
pigs fed diets containing AGP. They found an average optimal SID
Trp:Lys ratio of 18% when pigs were fed corn, pea, SBM diets
whereas the optimal ratio was 17% when corn, pea, barley, wheat
diets were fed. Eder et al. (2003) and Zhang et al. (2012) reported
the average SID Trp:Lys of 21% to optimize the ADG, FCR or N
retention of 25-50 kg crossbred pigs fed AGP-free diets. Similarly,
van der Aar et al. (2012), based on broken-line regression,
observed that the ADG, FCR and feed intake (FI) of 25-55 kg
crossbred gilts maximized at 20% SID Trp:Lys ratio.
For 55-110 kg crossbred gilts, Van der Aar et al. (2012)
estimated an optimal SID Trp:Lys ratio of 19% to maximize the
ADG, FCR and FI. Similarly, Guzik et al (2005) derived an optimal
SID Trp:Lys ratio of 19% to maximize the ADG and FCR of 80-105
kg barrows. Interestingly, when corn DDGS was included at 30%
in the diets, the average optimal SID Trp:Lys ratio of 19.5%
(Salyer et al., 2013) and 19.8% (Goncalves et al., 2015) were
estimated for 66-124 kg and 107-125 kg PIC pigs, respectively.
More recently, Xie et al. (2014), based on broken-line and
quadratic regressions, estimated 22% SID Trp:Lys as optimal for
ADG and FCR of 67-96 kg pigs fed low CP-AGP free diets. Eder
et al. (2003) derived an average SID Trp:Lys of 20% to optimize
the ADG, FCR and N retention of 80-115 kg crossbred pigs fed
AGP-free diets. Kendall et al. (2007) conducted 3 experiments to
determine the optimal Trp:Lys ratios for finishing pigs with the BW
ranges of 89 to 123 kg. They observed the best ADG and FCR
were achieved at the average SID Trp:Lys ratios of 15, 20 and
17%, respectively in these studies. More recently, Ma et al.
(2015b) reported the SID Trp:Lys ratio of 18% for 89-121 kg
crossbred pigs.
There is considerable variation in optimum dietary Trp:Lys
ratio among published data. These variations may be attributed to
differences in health status, statistical models, and the use of
AGPs and digestibility of Trp in feed ingredients used. Indeed, Le
Floc’h et al. (2007) reported that the optimal feed intake and ADG
of weaned pigs kept under poor sanitary conditions were achieved
at 21% Trp:Lys ratio whereas 18% was sufficient for pigs kept
under good sanitary conditions. Based on these results (Table 7),

the average optimal SID Trp:Lys ratio is 20% for 25-45 kg pigs and
19% for 80-115 kg pigs. These estimates are slightly higher than
the NRC (2012) recommendation of 17 and 18% for the
corresponding BW range.
OPTIMAL VALINE TO LYSINE RATIOS FOR GROWING PIGS
Leucine (Leu), isoleucine (Ile) and valine (Val) have a

unique branched-chain structure and are therefore commonly
referred to as branched-chain amino acids (BCAA), which have
important physiological functions especially in the regulation of
muscles protein synthesis. The metabolism of BCAA is unique and
different from other AA in that all three BCAA share the first two
steps of their catabolism and compete for the same degrading
enzymes. The BCAA antagonism and particularly the growth-
depressing effects of excess Leu are well known in several
species and a comprehensive review on this topic has been
previously described by Harper et al. (1984). Valine and Ile
become limiting after Lys, Thr, Met and Trp in low CP diets for pigs
(Liu et al., 2015). The overview on the published dietary Val:Lys
ratio estimates are given in Table 8.
Table 8 - Review of optimal SID Val:Lys ratios for growing-finishing

pigs
BW Sex1 Breed2 LysLeu:Lys Response Optimal SID Model4 Reference
kg (SID, %) criteria Val:Lys, %
13-32 M PIC 1.10 114 ADG, G:F 65 BL Gaines et al., 2011
21-33 B PIC 1.10 n/a ADG, G:F 66 BL Kendall et al., 2004
3
31-45 G PIC n/a n/a ADG 67 BL Goncalves et al., 2016
25-45 G PIC n/a n/a ADG 68 QBL Goncalves et al., 2016
26-46 M D×LR/LW 0.90 1.13 ADG, G:F 67 BL, QBL Liu et al., 2015
22-45 M D×LR/LW 0.83 130 ADG, G:F 69 BL Waguespack et al., 2012
25-40 67 Average
65 NRC, 2012
49-70 M D× LR/LW 0.73 123 ADG, G:F 69 BL, QBL Liu et al., 2015
71-92 M D× L /LW 0.61 128 ADG, G:F 70 BL, QBL Liu et al., 2015
94-119 M D× L /LW 0.51 141 ADG, G:F 70 BL, QBL Liu et al., 2015
70-95 70 Average
65 NRC, 2012
1
2
D = Duroc; LW = Large white, LR = Landrace.
3
4
Q = quadratic; QBL = quadratic broken-line; BL = linear broken-line.
Gaines et al. (2011) determined the optimal SID Val:Lys to be

65% in 13-32 kg pigs fed corn-SBM based diets. Based on ADG and
G:F responses, Kendall et al. (2004) reported the optimal SID Val:Lys
ratio to be 66% in 21-33 kg pigs. Recently, Goncalves et al. (2016)
conducted two dose response trials and based on ADG and G:F
responses, the optimal SID Val:Lys ratio was 68% in 25-46 kg PIC
gilts, and 67% in 26-46 kg PIC gilts, respectively. Waguespack et al.
(2012) reported that the ADG and G:F of were optimized at a SID
Val:Lys of 69% in 22-45 kg crossbred pigs. Liu et al. (2015) conducted
a series of 4 experiments to estimate optimal Val:Lys ratio for growing
and finishing pigs. Based on broken-line and quadratic broken-line
regressions of ADG and G:F responses, they estimated the average
optimal SID Val:Lys ratio of 67% for 26-46 kg, 69% for 49-70 kg, and
70% for both 71-92 kg and 94-1119 kg pigs.
Based on these results, the average optimal SID Val:Lys ratio
is 67% for 25-40 kg pigs and 70% for 70-95 kg pigs. These estimates
are slightly higher than the NRC (2012) recommendation of 65%.
OPTIMAL ISOLEUCINE TO LYSINE RATIOS FOR

GROWING PIGS
A brief summary on the optimal dietary Ile:Lys ratio

estimates are given in Table 9.
Table 9 - Review of optimal SID Ile:Lys ratios for growing-finishing

pigs
BW Sex1 Breed2 SDBC3 Lys Leu:Lys Response Optimal SID Model4 Reference
kg (SID, %) criteria Ile:Lys, %
15-30 G PxLW/LR 0 0.99 118 ADG, G:F 55 BL Castilha et al., 2012
20-45 M D× LR/LW 0 0.83 130 ADG, G:F 53 anova Waguespack et al., 2012
24-40 M PIC 4.5 0.91 129 ADG, G:F 54 BLQ, ExpoHtoo et al., 2014
20-40 53 Average
52 NRC, 2012
58-76 M PIC 0 0.80 123 ADG, G:F 56 BL, QBL Fu et al., 2005a
58-76 M PIC 7.2 0.80 200 ADG, G:F 60 BL, QBL Fu et al., 2005a
58-76 53 NRC, 2012
97-122 M PIC 0 0.52 148 ADG, G:F 53 BL, QBL Fu et al., 2005b
90-120 B PIC 3.9 0.52 238 ADG, G:F 60 BL, QBL Fu et al., 2005b
92-118 53 NRC, 2012
1
2
D = Duroc; LW = Large white, LR = Landrace, P = Pietrain.
3
SDBC = spray-dried blood cells.
4
Q = quadratic; QBL = quadratic broken-line; BL = linear broken-line, Expo = exponential.
In a dose response trial conducted by Castilha et al. (2012),

the optimal SID Ile:Lys was 55% based on the broken-line analysis
of ADG and G:F responses of 24-40 kg gilts. Waguespack et al.
(2012) also observed the greatest ADG and G:F of 20-45 kg
crossbred pigs at dietary SID Ile:Lys of 53%. Htoo et al. (2014)
also reported the optimal SID Ile:Lys ratio of 54% for 24-40 kg PIC
pigs based on ADG and G:F responses. From these studies, the
average optimal SID Ile:Lys ratio is 53% for approximately 20-40
kg pigs which are fed diets with normal Leu contents.
For 58-76 kg PIC pigs, Fu et al. (2005) reported that the
ADG and G:F were optimized at the SID Ile:Lys of 56% when
corn-SBM diets contained non-excess Leu (123% SID Leu:Lys).
However, the optimal SID Ile:Lys was estimated at 60% when
7.15% spray-dried blood cells (SDBC) was added to the corn-SBM

diets having an excess level of Leu (200% SID Leu:Lys).
In 97-122 kg pigs, Fu et al. (2005b) determined the SID
Ile:Lys to be 53% using corn-based diets (SID Leu:Lys of 148%).
However, they found in another trial that the optimal SID Ile:Lys
was increased to 60% when 3.9% SDBC was added to the corn-
based diets with excess SID Leu:Lys of 238% in 90-120 kg pigs.
The NRC (2012) recommends a dietary SID Ile:Lys of 52 to 53%
for growing and finishing pigs. Based on the literature review, the
optimal Ile ratio is not constant and increased when the diets
contain an excess level of Leu.
OPTIMAL LEU TO LYSINE RATIOS FOR GROWING PIGS
Research on the Leu requirement is almost non-existent,

mainly because Leu has been thought unlikely to be deficient in
typical pig diets. However, along with Val and Ile, it is meaningful
to have knowledge about the optimal Leu:Lys ratio to judge if the
dietary Leu level is at optimal or rather excess level. Recently,
Garcia et al. (2015), applying a linear broken-line regression,
reported that the ADG and FCR of 27-43 kg pigs were optimized
at a dietary SID Leu:Lys ratio of 104 and 109%, respectively,
resulting an average ratio of 107%. This estimate is in close
agreement with an early optimal Leu:Lys ratio of 110% for 25-50
kg growing pigs (Wang and Fuller, 1989) but slightly lower than
the NRC (2012) recommendation of 100%.
Table 10 - Review of optimal SID Leu:Lys ratios for growing pigs

BW Sex1 Breed2 Lys Ile:Lys Val:Lys Response Optimal SID Model3 Reference
kg (SID, %) criteria Leu:Lys, %
27-43 M LWxD 0.80 63 78 ADG, G:F 107 BL Garcia et al., 2015
100 NRC, 2012
1
M = mixed-sex.
2
D = Duroc; LW = Large white.
3
Q = quadratic; QBL = quadratic broken-line; BL = linear broken-line.
Based on the literature review, the requirement of Lys has

increased over time with increases in lean gain capacity of today’s
high lean pigs, i.e. approximately 20 g of SID lysine/kg of gain for
25-50 kg growing pigs and 21g of SID lysine/kg of gain for 50-115
kg finishing. The optimal dietary NE content may slightly decrease
for the high lean pigs due to a reduction in maintenance energy
need but more research is needed to better understand on energy
utilization. In response to the need of reliable ideal protein ratios
particularly in low protein-amino acids fortified diets, numerous
studies have been reported the optimal ratios of the other amino
acids. Based on the available literature, the optimal ratios of some
functional amino acids are higher than previously assumed optimal
values which may be attributed to differences in health status
associated with the ban of AGP in the diets, and the difference in
regression models used. Future research is warranted to
quantitatively estimate the increased need of functional amino
acids for maintaining optimal immune (health) status and
performance of pigs raised under sub-optimal conditions. More
research is also needed to estimate the requirement of net energy
in different pig diets.
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VALIDATION OF THE NUTRITIONAL REQUIREMENT
FOR HIGH PERFORMANCE SWINE
1 1 1 1
Uislei Orlando , Márcio Gonçalves , Wayne Cast , Matthew Culbertson ,
2
and Keysuke Muramatsu
1
PIC North America, 2JBS Foods - Brazil
100 Bluegrass Commons Blvd, Suite 2200, Hendersonville, TN 37075 (USA)
ABSTRACT
Genetic advancement continues to deliver outstanding

results at the commercial level with an accelerating rate of
improvement. The use of large-scale commercial nutrition trials
allows us to determine nutrient specifications to maximize the
profitability for swine producers worldwide. This article will cover
the most important topics to be taken into consideration in the
process of validating and applying nutritional requirement for high
potential genetic.
INTRODUCTION
Reproductive and growth performance traits have made

great strides in the last decade through genetic improvement. The
exponential increase in availability of cutting-edge genetic
technologies, such as genomics, is continuing to accelerate the
rates of genetic improvement. In addition, large-scale commercial
research allows us to capture this genetic potential and increase
profitability for swine producers around the globe. The increase in
feed cost in the last decade has prompted swine producers to
continue to focus on whole-herd feed efficiency. Thus, with the
improved rate of gain and feed efficiency, a tailored requirement
for each pig production system should be used based on market
scenarios and ingredient prices.
Finally, before applying new nutritional requirements or
design a trial to determine requirement is necessary to understand
about the goals or market strategies (e.g. fixed time or fixed
weight, payment on a carcass or live basis), limitations from the

production system (e.g. phase feeding, health challenges, feeder
type and space, stocking density) or fluctuation of market prices
(Figure 1) and cost of production (Figure 2).
Source: Hurley and Associates - Hog review – 31/12/2016,

http://www.hurleyandassociates.com
Figure 1 - Weekly national hog price in the USA.

Source: Hurley and Associates - Hog review – 31/12/2016,

http://www.hurleyandassociates.com
Figure 2 - Nearby weekly corn, Soybean meal and DDGS prices

and DDGs/corn ratio in the USA.
GENETIC IMPROVEMENT
In 2006, the average number of weaned pigs per mated

female per year was 21.5 in the United States (Pig Champ, 2006).
Today, the top 10% of the farms are above 30 weaned pigs per
mated female per year, with an average of 26.2 (PIC internal
database; 700,000 sows). With the use of relationship-based
selection using genomics, the rate of progress of genetic selection
of PIC pigs has increased by over 35% beginning in 2014 (Figure
3) and is starting to be realized by customers this year as the
rollover of the replacement gilts occurs through the multiplication
and commercial flows. With the use of this and other new
technologies in the pipeline, it can be suggested that by 2062 a

commercial female might be expected to wean 56 pigs per year.
Source: Orlando et al. (2016)
Figure 3 - Accelerating progress of genetic improvement.
Table 1 shows the rates of changes of genetic improvement

for different traits of PIC pigs in the last 5, 3, and 1 year
Table 1 - Rates of changes of genetic improvement for different

traits of PIC pigs in the last 5, 3, and 1 year
5 Year Avg. 3 Year Avg. 1 Year Avg.

Index 13.2 16.0 19.5
Pigs weaned/sow/year 0.8 0.9 1.1
Kilos weaned/sow/year 5.4 5.94 8.03
Pigs marketed/sow/year 0.8 0.9 1.0
Total kilos marketed/sow/year 129.8 144.8 186.3
Feed conversion -0.01 -0.02 -0.02
Profit per pig, $ / pig 2.63 3.20 3.89
Source: Orlando et al., (2016).
Increasing prolificacy, if not balanced for potentially

associated negative impacts, may increase the incidence of
lightweight low quality piglets. PIC has long selected for quality
weaned pigs with superior growth performance and has increased
the focus on this area by including individual piglet birth weight
(Figure 4) in the genetic program (along with total born, stillborn,
pre-weaning mortality, and weaning weight). There was a
reduction in individual piglet birth weight of about 100 g from 2006
to 2013, and this reduction seems to be fully reverted now at the
genetic nucleus level. This type of program is believed to allow for
an increase on the number and weight of weaned pigs/sow/year
and maintain exceptional growing pig performance.
Source: Orlando et al., (2016)
Figure 4 - Trend in genetic improvement in birth weight and total

born at PIC genetic nucleus.
Innovation model
To better understand what validation means or why

companies invest one of the most important assets “time” and also
a considerable amount of money in this area, we need to start by
describing the innovation model. Although the majority of
professionals involved in applied new technologies consider the
process linear and simple. As soon as an opportunity is identified
and applied the production system will capture the potential
savings. However, very often are neglected some intermediate
steps such as experimental and validation before applied thus
capture the potential savings (Figure 5).
Source: Orlando 2016
Figure 5 - Innovation model in animal nutrition in the integrator

company.
Research and development in agricultural chain is different

than the historical one based in the transfer of knowledge from the
basic science (academy) to practice (field) as described in the
Figure 6 (Leaver, 2010).
Figure 6 - Historical model of research and development chain.

Adapted from Leaver, 2010.
As Leaver (2010) describes, there are many other points to

be considered innovation for agriculture. This model is not a simple
top down flow, as many professionals believe. Likewise,
considering the private funding as fundamental to improve the
innovation process and avoid lack of innovation with applied level
(Figure 7).
Figure 7 - Agricultural research and development chain: Academic

and U.K. model. Adapted from Leaver, 2010.
There are three main research models: based in the

university, private companies (e.g. progressive companies,
nutrition companies and integrators) and mixed model based in
partnership between universities and private companies. Although
it is not clear-cut which approach is the best, it is possible to state
pros and cons of each approach. While the university model is
more focused in basic science and pure innovation, the private
model is focused in large-scale experiments those represent more
the field conditions, thus, reducing the time between research and
application. Oftentimes the private model is limited in ideas and
lack of creativity, and not open to peer-review and result sharing.
The mixed model is interesting because joins mutual interests and
fix some of the weakness from each individual model (Figure 8).
Figure 8 - U.S. Swine applied research and development models.

Adapted from Tokach et al., (2010).
VALIDATION AND APPLICATION OF RESEARCH IN LARGE

PRODUCTION SYSTEMS
The use of large-scale commercial nutrition trials allows the

nutritionist to determine nutrient specifications to maximize
profitability of the system. Furthermore, increase confidence in the
results for faster application. Although the large-scale is the best fit
to validate and apply innovation, it is important take into
consideration has pros and cons (Table 2).
Table 2 - Advantages and disadvantages between small and large

scale research facilities
Model Advantages Disadvantages

 Low risks involved  High cost relative
Small-scale  Basic science  Low replicability (field)
 Low pressure for results  Low apply focus
 Low cost (operation)  Hard to share results
 Strong replicability  Confounding facts
 Data controlling  Prioritizing/time
 Possibility to work with  Struggle to have many
Large-Scale
closeouts as reps treats
 Fast decisions  High risk (losses)
 Trainings high skills  Comparisons not
professional in the field contemporaneous
Large-scale nursery and finishing nutrition trials with

automatic feeding system
The concept is based on having one scale that is the size of

pens (i.e., 25 pigs per pen) with an automatic feeding system in a
commercial facility. This will yield results that are readily applicable
to the rest of the system due to similar production challenges such
as stocking density, quality of stockmanship, health challenges,
and feeder space.
Electronic Sow Feeding (ESF) system
The current growing concern with animal welfare increase

the importance of food producing companies in South and North
America claims they change the production system next soon. It
means that sows will be group-housed instead of individually
housed. Although this transition has a high investment cost it
requires the correct management practices to achieve similar
results to the traditional production system.
The increased number of farms with ESF increased the
opportunities for gestation sow nutrition research.
Two hoppers over the Large-scale commercial research barn

station feeds
In a recent trial conducted in a modern 5,600 commercial

ESF sow farm with automatic weighing of the sows Thomas (2017)
collected data from a population of sows and the number of total
weight observations was over 660,000. The collection of weight
and intake data, allows to document average daily gain, average
feed intake, and feed efficiency of gestating PIC sows (Figure 9).
Source: Thomas, (2017)
Figure 9 - Data Management: Body Weight.
Precision nutrition
This concept is based in feeding to the requirement of each

individual and is believed by some researchers to improve nutrient
utilization; however, validation in large-scale trials and evaluation
in the production systems is a necessary step before final
application.
Source: Research barn at Unesp Jaboticabal Campus
Pen scale concept
Wean-to-carcass data collection
In some experiments, pigs are tagged with RFID ear tags

and tracked individually at packing plants to capture carcass
information. This technology allows researchers to create strong
models with more precision and evaluate economical requirements
based in carcass or even primal cuts.
Kinect depth sensor
The use of kinect cameras to estimate individual or group

body weight seems very promising for real-time weight estimation
with great accuracy (97.5% at group level with error of 820 g and
96.2% at the individual level with error of 1.23 kg; Kashiha et al.,
(2014). Overall R2 was 0.975 with standard error of 0.0182 (Figure
10).
Figure 10 - Observed versus estimated weights over six days of

four pens with ten pigs per pen (240 data points).
Source: Lee et al. (2016)

Figure 11 - Pig housing unit installed with a stationary Kinect sensor.
This concept is in the very beginning and must be improved

before application; however, likelihood of application seems high
both from a production and from research standpoint.
DECISION MAKING IN DIET FORMULATION
From a macro level, once growth and feed intake in the

specific production system are known, the first step in diet
formulation is to define the most economical net energy (NE) level.
The second step is determining the standardized ileal digestible
(SID) lysine (Lys) dietary concentration based on the SID Lys: NE
ratio. Next, the other SID amino acids (AA) are defined as a ratio
to SID Lys. Finally, the levels of macro minerals, trace minerals
and vitamins are defined to achieve the requirement in amount of
nutrients (i.e., grams, milligrams, or International Units) per pig per
day.
The economic implications of fixed time vs. fixed weight
A key concept to consider when formulating diets for a

specific production system is to understand if the system is
marketing pigs on a fixed time or a fixed weight basis. Fixed time
means that the system does not have extra or flexible space in the
production flow. For example, when a finishing barn reaches 120
days of placement, the pigs are marketed and the barn is emptied
for the next group of pigs. Fixed time can also be explained as
being space short and fixed weight as space long. Fixed weight
program, however, means that the system has some flexible
amount of space available in the production flow and, thus, pigs
can be left in the barn until they reach a target weight optimum for
the given carcass value payment structure of the processing plant.
The difference between these two scenarios is important because
it changes the relative value of growth rate. The value of weight
gain in a fixed time system is more valuable given the fixed
constraint on number of growing days available; however, in the
fixed weight system, pigs can stay in the barn at a fixed space cost
(i.e., $0.11/pig/day) and, therefore, the economic value of weight
gain by a given nutritional or management strategy is smaller
compared to a fixed time scenario. Production systems will often
be on a fixed weight basis during winter when pigs are growing at
a faster rate and on a fixed time basis during summer when pigs
are growing at a slower rate. The important point is that these two
scenarios represent the range of economic optimums and
evaluating both scenarios can be an effective tool for evaluating
economic sensitivity of dietary changes.
The concept of optimum nutrient levels to maximize
profitability in a fixed time program relative to fixed weight scenario
is illustrated in Figure 12. Tryptophan (Trp) to Lys ratio can have a
significant impact on growth rate. In this specific scenario, varying
tryptophan to lysine ratio has a much larger economic impact on a
fixed time system than a fixed weight system simply because
weight gain offers a greater marginal economic return compared to
the fixed weight scenario. For additional information on the value
of alternative Trp to Lys ratios, please visit
http://www.lysine.com/en/tech-info/TrpLys.aspx to download a free
dynamic economic calculator for the most economic Trp to Lys
ratio specific to a production system.
Figure 12 - Standardized ileal digestible tryptophan: lysine ratio for

maximum profit on a fixed time and fixed weight basis
(PIC 337 × 1050; Kansas State University and
Ajinomoto Heartland, 2016).
STRATEGIES FOR DIET FORMULATION
There are multiple strategies, or targets, that are commonly used

for diet formulation. Some of the commonly used approaches are:
 Growth performance basis

o Maximize average daily gain (ADG)
o Minimize feed efficiency (F/G)
 Cost reduction basis
o Minimize cost per kg of diet
o Minimize feed cost per kg of gain
 Profit maximization basis
o Maximize income over feed cost (IOFC)
o Maximize income over feed and facility costs
(IOFFC)
o Maximize income over total cost (live or
carcass)
A summary showing how these targets can impact

formulation strategies and the resulting diets is shown in Figure 13.
These results show the levels of SID Lys to optimize the different
strategies listed above. Note that the SID Lys level to maximize
profit is greater than that to minimize cost. The economic optimum
SID Lys level is dynamic and depends on the market prices. Each
of these concepts, and some of the relative risks and rewards, are
explained below in more detail.
Figure 13 - Example of levels of standardized ileal digestible

lysine to optimize different outcomes for PIC pigs (20-
to 25-kg pig; PIC internal data).
Formulating for maximum performance
The SID Lysine level to improve F/G is generally greater

than that to maximize ADG. However, formulation targeting
maximum performance does not take into account any economic
measurement but only considers the impact on the biological
response.
Formulating for minimum cost
To minimize the diet cost, the nutritionists set the nutrient levels
needed and use a least cost formulation software to achieve the
minimum diet cost possible but still meet the needed requirements.
Thus, diet cost is technically an economic variable; however, it
does not account for any changes in performance. Feed cost per kg of
gain is calculated by multiplying F/G by the cost per kg of feed and,
therefore, feed cost per kg of gain takes into account F/G. However,
this approach does not take into account any changes in ADG, pig
price, or the cost of each extra day in the barn.
Feed cost per kg gain =
Formulating for maximum profit
Income over feed cost (IOFC), on the other hand, takes into
account the market price and the value of weight gain under a
fixed time scenario:
IOFC = (market price per kg live weight weight

gain)
Income over feed and facility costs (IOFFC) is similar to

IOFC, however it is suitable for a fixed weight scenario:
IOFFC = (market price per kg live weight weight gain) - (feed cost
per kg gain x weight gain) – (cost per pig space x days in the phase)
Putting it all together
The formulation concept of feed cost per kg of gain

generally leads to the conclusion of cheaper diets; however, often
that is not necessarily the optimum level to maximize net profit.
Income over total cost (IOTC) takes into account the dilution effect
of the extra gain over each kg of live or carcass produced. For

example, let us assume that the cost of the weaned pig was $40.
Therefore, a production system with 121 kg of gain from weaning
to market results in a cost of $0.3306 per kg that will be related to
the cost from the weaned pig. However, if a given nutritional or
management strategy increases the weight gain to 123 kg, the
cost per kg related to that initial weaned pig cost will change to
$0.325 or 1.7% reduction in cost.
To calculate income per kg of live weight produced:
IOTCL = [market price per kg - ((1/market weight) x (feed cost per

pig + other costs per pig + feeder pig cost))]
Or to calculate income per kg of carcass weight produced:
IOTCC = [market price per kg - ((1/market weight/% yield) x (feed

cost per pig + other costs per pig + feeder pig cost))]
The following examples use these principles for comparison

of a few specific scenarios and the impact on income over feed
cost and income over total cost on a carcass basis:
Comparison of minimizing cost vs. maximizing profit per pig
Table 3 - Scenarios and assumptions

a
Scenario 1 Scenario 2
Assumptions Fixed time/no added fat diet Fixed time/3% added fat diet
ADG, kg 0.816 0.841
F:G 2.800 2.632
Days on feed 112 112
b
Diet cost, $/kg 0.229 0.245
a
Assuming each 1% added fat improves gain by 1% and F: G by 2%. This response
can vary from system to system and by season.
b
Assuming costs of soybean meal, corn, and choice white grease at $350/ton,
$3.60/bushel (25.5 kg), and $0.68/kg, respectively.
Diet cost should have manufacturing and delivery included and

not just ingredient cost because this is a more accurate reflection of
the cost of the feed consumed and the value of the performance
differences.
Calculations
Scenario 1 (no added fat): 112 days x 0.816 ADG = 91.4 kg gain in
the finishing
Feed cost per pig = 91.4 kg gain x 2.80 F/G x $0.229 feed cost/kg =
$58.60
Scenario 2 (3% added fat): 112 days x 0.841 ADG = 94.2 kg gain
in the finishing
Feed cost per pig = 94.2 kg gain x 2.632 F/G x $0.245 feed cost/kg
= $60.74
In conclusion, the feed cost per pig in scenario 2 is $2.14 greater

than scenario 1.
Thus, scenario 1 has the lowest feed per cost per pig;
However, in scenario 2 there are more kg produced per pig. Thus,

this needs to be taken into consideration:
Considering the market pig price equal $1.21/kg and recalculating

using IOFC:
Scenario 1:
IOFC (Sc1) = ($1.21 pig price/kg x 91.4 kg gain) – ($58.60 feed cost
per pig) = $51.99 per pig
IOFC (Sc2) = ($1.21 pig price/kg x 94.2 kg gain) – ($60.74 feed cost
per pig) = $53.24 per pig
In conclusion, the income over feed cost per pig in the

scenario 2 is $ 1.25 better than scenario 1, thus, adding fat in this
scenario is more profitable.
Income over total cost
Assumptions
Carcass yield = 74%
Carcass price = $1.65/kg
Feeder pig cost (22.7 kg) = $55
Other costs (facilities/transport/medicines/vaccines/slaughter) =
$14.56 per pig
Calculations on a live basis
IOTCL sc1 = [$1.21- ((1/(22.7+91.4)) x ($58.60+$14.56+$55.0))] =

$0.0868 per kg live weight produced
IOTCL sc2 =[$1.21- ((1/(22.7+94.2)) x ($60.74+$14.56+$55.0))] =
$0.0954 per kg live weight produced
Scenario 2 (3% added fat) is 9.9% ($8.6/ton of live weight) more

profitable than 1 (no added fat) in this market situation on a live
basis.
Calculations on a carcass basis
IOTCC sc1 = [$1.65 - ((1/(22.7+91.4)/0.74) x ($58.60+$14.56+$55.0))] =

$0.1321 per kg carcass weight produced
IOTCC sc2 = [$1.65 - ((1/(22.7+94.2)/0.74) x ($60.74+$14.56+$55.0))] =

$0.1437 per kg carcass weight produced
Thus, scenario 2 (3% added fat) is 8.8% ($11.62/ton of carcass

weight) more profitable than 1 (no added fat) in this simulation.
A summary of absolute and relative economic differences

between scenarios are presented in Table 4.
Table 4 - Absolute and relative economic differences between

scenarios 1 and 2
Differences (Scenario 2 – Scenario 1)

Absolute Relative (%)
Diet cost, $/kg 0.016 + 7.0%
Feed cost per pig, $/pig 2.14 + 3.6%
Feed cost per kg produced, $/kg 0.004 + 0.6%
IOFC, $/pig 1.25 + 2.3%
IOTC Live weight based, $/ton 8.60 + 9.9%
IOTC carcass based, $/ton 11.62 + 8.8%
In conclusion, there are multiple strategies and approaches

for diet formulation. It is important to use an approach that takes
into account the value of performance (i.e., ADG, F/G, and yield)
but also the fixed time or fixed weight nature of the system.
Therefore, using approaches such as income over feed (and
facility) costs or income over total cost on a carcass basis are
suitable solutions to robustly maximize the profitability of swine
operations.
NUTRIENT REQUIREMENTS OF PIC PIGS
Last year the global PIC nutrition team launched the PIC
2016 nutrition manual with an update on nutrient specifications for
different phases of production. The requirements are based in the
latest 50 large-scale scientifically designed experiments with over
60,000 PIC pigs.
The manual was built up in three sections that lays out the
fundamentals of our nutrition recommendation: first, it summarizes
the principles of diet formulation; secondly, it lays out how different
nutritional components can help fulfill those dietary formulation
principles; and then it details how the basic diets vary for pigs
depending on phase of production.
After these sections that lay out the fundamentals of our nutrition
recommendation, we have included some tools and deep dives in
specific topics that will help you optimize the diets for your pigs.
Finally, you find the nutrient specification tables that you can
use to optimize your diets for successful nutrition of PIC pigs.
Recommendations are based on published research, PIC internal
research, research from universities, and commercial large scale
designed experiments. The nutrient specifications have been
validated in commercial environments. The National Swine
Nutrition Guide (2010) and National Research Council (2012)
publications serve as the basis for certain information. Concepts
and the basis for recommendations are discussed in greater detail
in other technical memos.
The PIC nutrition manual is a dynamic manual. PIC will
continue to update this manual as new research becomes
available and share them with you through nutrition updates and
the PIC website. Access
http://na.picgenus.com/enewsletter_sign_up.aspx to sign up.
Breeding herd nutrition and feed efficiency
The breeding herd represents 10 to 15% of the total feed

cost of a market pig. A series of recent research with PIC sows
during late gestation has shown that the increase of feed in late
gestation has modest (40 g) impact on individual birth weight of
piglets from gilts and little to no impact on birth weight of litters
from sows (Shelton et al., 2009; Soto et al., 2011; Ampaire and
Levesque, 2016; Greiner et al., 2016; Gonçalves et al., 2016a).
There has also been documented in a large-scale study an
increase in stillborn rate in litters from sows that were fed 6.75
Mcal NE/d instead of 4.50 Mcal NE/d in late gestation (Gonçalves
et al., 2016a). Therefore, it is currently recommended to only
increase the amount of feed in late gestation to gilts, but not sows.
It is recommended that gilts during gestation are fed 4.50 Mcal

NE/d until d 90 of gestation and then 6.75 Mcal NE/d, whereas for
sows it is recommended to feed 5.4 Mcal NE/d until d 28 and 4.50
Mcal NE/d afterwards until transferring the sow to the farrowing
house. This demonstrates the efficiency of PIC females and
minimizes the amount of feed per sow per year as well as the
amount of breeding herd feed per weaned pig.
Lysine requirement update
PIC has delivered dramatic increases in lean gain and

efficiency to swine producers around the globe in the last decade.
With an increase in growth rate and improved feed efficiency it is
expected that, overtime, the nutrient concentration of the diets
need to be updated to match the pig’s needs to achieve its genetic
potential. Lysine is, typically, the first limiting amino acid in swine
diets.
Thus, a meta-analysis was to evaluate the standardized
ileal digestible lysine requirement for PIC pigs. To accomplish this
objective, a total of 27 commercial experiments were used in the
meta-analysis with a total of 45,102 pigs. Nine out of the 27
experiments were in partnership with JBS United. Each treatment
within an experiment was considered as an observation (n=213)
and each experiment was used as random effect. All dam lines
were Camborough 1050. Ingredient loadings values used were
based on NRC (2012). Requirement estimation models were
implemented as per Gonçalves et al. (2016b). Models that differed
in their bayesian information criterion values by at least 2 points
were considered to have meaningful differences in their data fit
(Raftery, 1996). If more than one model had adequate fit, then the
average requirement estimate was used. The requirements
presented are an average of the requirement for ADG and F/G.
Requirements estimates for boars were based on relative
differences from barrows published by Bertram et al., (2014) and
NRC (2012).
Due to the high potential for protein deposition, adequate

amino acid supply is important for successful production of PIC
pigs. Amino acid deficient diets can have negative influences on
growth performance and behavior (Fraser et al., 1991; NRC,
2012).
The Figure 14 show the relative changes in Lys requirement
from 2008 versus 2016 meta-analyses in a corn-soybean meal
based diet. Figure 15 show Lys requirement for barrows, gilts, and
boars in a metabolizable energy (ME) basis. Finally, the Lys to
calorie ratio equations for gilts, barrows, and boars are presented
in metric unit systems in Tables 5.
Figure 14 - Example of standardized ileal digestible (SID) Lysine

(Lys) for PIC finishing pigs from 2008 and 2016 meta-
analyses for a corn/soybean meal based diet (average
of barrows and gilts) with 3.31 Mcal ME/kg (2.44 Mcal
NE/kg).
Figure 15 - Standardized Ileal Digestible (SID) Lysine (Lys) to net

energy (ME) ratio for different body weight and gender.
Table 5 - Standardized Ileal Digestible (SID) Lysine (Lys) to

metabolizable (ME) and net energy (NE) ratio
equations for PIC pigsa,b,c
Gilts
2
= 0.000043 × (body weight, kg × 2.2046) –
SID Lysine:Calorie ME 0.02154 × (body weight, kg × 2.2046) + 4.9538
2
= 0.000056 × (body weight, kg × 2.2046) –
SID Lysine:Calorie NE 0.02844 × (body weight, kg × 2.2046) + 6.6391
Barrows
2
= 0.000031 × (body weight, kg × 2.2046) –
2
= 0.000042 × (body weight, kg × 2.2046) –
Boars
2
= 0.000034 × (body weight, kg × 2.2046) –
2
= 0.000046 × (body weight, kg × 2.2046) –
a
These specifications should be used as a guide. They require adjustment for
feed intake, local conditions, and markets.
b
Lysine specifications are based on a series of 27 trials conducted under
commercial research conditions (9 of them in partnership with JBS United).
These equations are only valid for pigs from 23 to 135 kg BW.
c
Requirements estimates for boars were based on relative differences from
barrows published by Bertram et al., (2014) and NRC (2012).
Understanding the biology and economics of PIC pigs
PIC has recently published the first lysine calculator based

in biological and economic requirements. The aim is to provide a
tool to help customers globally to make the best decisions while
creating their nutritional program for PIC pigs by tailoring to their
own production system while considering the volatility in the cost of
raw materials and market pig prices (Figure 16).
Figure 16 - Lysine biological and economic calculator of PIC pigs.
Both are available at:

http://na.picgenus.com/tech_support/nutrition.aspx
In conclusion, lysine requirements for PIC pigs are virtually the
same on an amount of grams of amino acids per unit of gain over
the last several years. However, they are greater than in the past
on a dietary concentration basis. This is probably due to increased
rate of growth and improved feed efficiency from modern lines.
Although it is important for practical reasons present the
requirement in a percentage basis, nutrients should be considered
in grams of intake per day based on the farm-specific feed intake.
PIC made great strides in the last decade improving

reproductive and growth performance traits, even more in the last
couple of years after the introduction of relationship-based
genomic selection which allowed an acceleration of the genetic
improvement. In addition, large-scale commercial nutrition
research has allowed nutritionists to capture this genetic potential
and increase profitability for swine producers around the globe.
The knowledge about not only the biology but also the
economics of feeding PIC pigs is fundamental to make more
powerful decision making, especially with the high volatility of cost
of raw materials and market pig prices observed in many regions.
Before validating and applying the new requirement
updates, the nutritionist must to know their own production
systems and its constraints and take into consideration the
biological and economic models that help achieve system goals by
using the maximum income over total cost.
Investments in cutting-edge technologies are important to
increase efficiency of production systems in this increasingly
competitive global swine industry. Partnership among private
companies, production system, and universities are fundamental to
capture value from new technologies, by improving the production
process and aiding risk management.
Finally, updating nutrient requirements based on
commercially scientifically designed experiments is of extreme
importance for replicability and robustness when applying the
generated knowledge. This can be achieved by using mixed
models that involves researchers from production companies and
universities.
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gestation on wean pig growth performance. J. Anim. Sci. 94:121. (Abstr.)
Bertram, M., Tokach, M., Pollmann, S., Nelson, D. 2014. Nutritional Guidelines for
Immunologically Castrated Male Pigs. Technical bulletin Zoetis. p. 8.
Fraser, D., Bernon, D.E. and Ball, R.O., 1991. Enhanced attraction to blood by pigs
with inadequate dietary protein supplementation. Can. J. Anim. Sci., 71:611-619.
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dose-responses to tryptophan and valine in finishing pigs. PhD Diss. Kansas
State Univ., Manhattan, KS.
Gonçalves, M. A. D., K. M. Gourley, S. S. Dritz, M. D. Tokach, N. M. Bello, J. M.
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acids and energy intake during late gestation of high-performing gilts and sows
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Woodworth, and R. D. Goodband. 2016b. An update on modeling dose–
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Sci. 94:1940-1950.
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increasing lysine or feed amounts in late gestation on piglet birth weights. J.
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Kashiha, M., C. Bahr, S. Ott, C. P. H. Moons, T.A. Niewold, F.O. Odberg and D.
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Nelssen, and S. S. Dritz. 2009. Effects of increasing feeding level during late
gestation on sow and litter performance. In: Kansas State University. Agricultural
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NUTRITIONAL REQUIREMENTS OF AMINO ACIDS
FOR PIGLETS
Etienne Corrent and Aude Simongiovanni
Ajinomoto Eurolysine S.A.S.
153 rue de Courcelles, 75017 Paris (FRANCE)
ABSTRACT
The increasing availability of crystalline amino acids (AA)

has definitely changed the piglet feed structure and the way to
deal with the nitrogen nutrition issue for these animals. It is
possible today to move from a formulation based on minimum
dietary crude protein to a formulation based on each indispensable
AA. This increasing practice results in a drastic reduction of the
dietary crude protein levels with the improvement of nitrogen
efficiency and health status of piglets without altering performance.
In these formulas, 7 AA are co-limiting and ideal AA profile must
be at its balanced level: no lack and no excess. Indeed, imbalance
between AA (like Branched Chain AA) can lead for instance in
feed intake reduction. The increasing knowledge about AA allows
to recommend a robust ideal AA profile that supports feed intake,
growth, nutrients efficiency, and health challenges : Lys (100), Thr
(65), Met+Cys (60), Trp (22), Val (70), Ile (53), Leu (100), His (32)
and Phe+Tyr (95). Review of trials using this AA profile shows
efficient results in terms of dietary protein reduction with
maintained performance. Although an ideal AA profile gives fixed
values, response to an AA can vary depending on the conditions
and more studies are needed to refine the impact of external
factors on an AA effect.
INTRODUCTION
Dietary protein reduction in swine is driven by economic,

environmental and societal issues - the three pillars of
sustainability. Formulating on the base of each indispensable
amino acids (AA) instead of a minimum crude protein (CP) level
allows for instance a reduction in feed cost, a decreased
dependency on soybean meal, and lower pressure on animal
health. Beside, the increasing availability of feed grade AA (i.e. L-
Valine) has made possible the further decrease in dietary CP and
changed the way to address risk management in feeds for
monogastric animals. Indeed, moving from a dietary formulation
based on protein to a more precise formulation that gives value to
each single AA is a shift from an unpredictable risk approach to a
controlled risk management. Formulating and relying on the
protein criteria (defined as nitrogen x 6.25) is only considering the
quantity of supply of various nitrogenous components but not their
quality nor the added value created by their interactions. In
contrast to nitrogen, AA are predictors of performance and the
precise control of the AA levels is crucial for performance and
profitability. It also gives more opportunity for innovative nutritional
choice and ends in the most economical feed solution by a better
adjustment of the feed recipe to the AA requirements. Increasing
knowledge about AA nutrition and focusing on their metabolic
roles is therefore mandatory. Focusing on AA, facts are that when
dietary CP is reduced, all quantitative AA concentrations are
decreased with variable impact on the supply of these AA, and
interactions between groups of AA arise. If minimum levels are
generally applied to the very first most limiting AA, the other
indispensable AA (IAA) are often not under control. The
information about IAA is also quite variable due to interactions (i.e.
Thr or Trp and health, Val, Ile and Leu antagonisms) or to protocol
issues. All the dispensable AA (DAA) are also decreased as
protein is decreased. The DAA levels are not controlled in
formulation while the literature provides new insights on their
essentiality even if the data are still scarce, variable and

sometimes contradictory. In this review, the focus is made on the
recent advances concerning Trp, Val, Ile, Leu, His, Phe, Tyr and
the dietary protein reduction in piglets. Data about Lys, Met and
Thr are based on previous works (van Milgen and Le Bellego
2003, Sève 1994).
AMINO ACIDS IN PIGLETS
From a pool of 20 AA, pigs can synthesize all the proteins

needed to fulfill their maintenance and growth requirements. Each
of the 20 AA is encoded in the DNA as codons, which enables the
protein synthesis. That is why the 20 AA can be considered as
essential for the protein synthesis during growth but 9 of them in
pigs are either not de novo synthesized at all or only in small
quantities. In pigs, Lys, Thr, Trp, Met (+Cys), Val, Ile, Leu, His,
Phe (+Tyr) are considered as IAA and must therefore be dietary
supplied. In suckling piglets, Arg is also considered as an IAA,
Table 1.
Table 1 - Usual classification of the 20 amino acids encoded

directly by the genetic code for pigs
Essential Amino Acids (AA) : AA involved in the biological functions that are
necessary for life supplied through the diet or synthesised de novo by animals
Indispensable AA Semi-dispensable AA Dispensable AA
AA that cannot be AA that can be synthesised de novo

AA that can be
synthesised de novo or but could become indispensable in
synthesised de
at a sufficient rate to specific situations when the synthesis
novo by the animal
maintain associated is not sufficient to cover the
in a sufficient rate
biological functions requirement
Lysine (Lys) Glycine (Gly)

Threonine (Thr) Serine (Ser)
Methionine (Met) Cystine (Cys) Proline (Pro)
Tryptophan (Trp) Alanine (Ala)
Valine (Val) Aspartate (Asp)
Isoleucine (Ile) Asparagine (Asn)
Leucine (Leu) Glutamine (Gln) Glutamate (Glu)
Histidine (His)
Phenylalanine (Phe) Tyrosine (Tyr)
Arginine (Arg)
With the feed-use AA available on the market it is possible

to formulate diets without minimum constraint on the CP level in
which at least 7 AA are co-limiting: Lys, Thr, Trp, Met (and Cys),
Val and the next one which determines the resulting dietary CP
level (Figure 1). Thus, determining AA requirements aims also at
identifying the next limiting AA in local situation, which then
depends on the local feedstuffs used.
Figure 1 - Example of the 7 co-limiting AA in a piglet formula in

which dietary crude protein is reduced, taking into
account requirement of IAA, and using feed-grade AA.
Ideal AA profile is the most practical tool to express the AA

requirements of animals. All IAA are expressed in ratio to Lys in a
digestible value. Specific protocols must be implemented to
estimate AA in ratio to Lys. The dose-response studies must use
basal diets with a sublimiting Lys level, otherwise the ratio would
not be valid and cannot be used in practical feeds (Boisen, 2003).
When sufficient number of dose-responses exist, meta-analysis
studies help in determining the general response to a specific AA
and in understanding the variability.
TRYPTOPHAN REQUIREMENT IN PIGLETS
Besides being a constituent of body protein, Trp also plays

other important roles in metabolism. It is involved in feed intake
regulation, in the immune response and in the animal’s defence
system. Increasing the Trp content in the diet has also been
shown to limit the impact of an unfavourable health environment
on performance in piglets. Being an IAA for pigs, Trp has to be
supplied by the diet in sufficient quantities to cover the animal’s
requirement for both growth and health challenged situations.
More than 130 studies about the effect of Trp on piglets’
performance have been listed by Simongiovanni et al. (2012) who
performed a meta-analysis concerning the piglet response to Trp.
This AA has been extensively studied due to a practical interest
linked to its numerous biological functions and its importance for
piglet nutrition. Among the 130 studies, 37 have been selected by
Simongiovanni et al. (2012) to assess a Trp requirement as a ratio
to Lys. Using a curvilinear-plateau model, and based on INRA
tables (Sauvant et al., 2004), requirement estimates for ADG,
ADFI and G:F have been determined varying from 20 to 22%
depending on the criteria to maximize. New results are in line with
the model estimated in the meta-analysis (Figure 2).
Figure 2 - Effect of Trp on the average daily gain (ADG) of piglets.

Comparison of the model determined by Simongiovanni
et al. (2012) with external Trp dose-reponses (not used
in the meta-anlaysis work).
VALINE REQUIREMENT IN PIGLETS
Valine is the fifth limiting AA after Trp in diets for piglets. Valine
is an AA for body protein deposition and growth, and a dietary
deficiency in Val affects the utilization of previous limiting AA and
consequently animal growth. Valine is a branched-chain amino acid
(BCAA), together with isoleucine and leucine. Due to their common
catabolic pathway, some interactions exist between them. It is
therefore very important to well know their requirements and to ensure
that the feed supplies a balanced BCAA profile.
With about 20 publications mostly in piglets, the number of Val
dose-response studies is lower than for Trp. Using most of the Val
dose-responses studies available, van Milgen et al. (2013) determined
by a meta-analysis work that there is a response of +5% for the ADG
when SID Val:Lys is increased from 64 to 69%. A data set of 12 trials

is presented in Figure 3. These trials were selected to allow to express
the performance according to dietary Val: Lys levels. It shows that on
average, a value of 70% SID Val: Lys enables to obtain the optimal
performance.
Figure 3 - SID Val: Lys dose-responses in piglets and meta-

analysis. Effect on average daily gain (ADG).
IMPACT OF THE BRANCHED-CHAIN AA METABOLISM ON

PERFORMANCE
Since AA cannot be stored, any excess of AA is

catabolized. The group of the BCAA (Val, Ile and Leu) shares in
common the two first steps of their catabolism (Figure 4). The
second step is under the influence of Leu which activates the
BCKDH enzyme complex leading to the catabolism of Val and Ile
when Leu is in excess. The Val and Ile catabolism is enhanced
even if these AA are deficient in the diet. Indeed, it has been
shown that when dietary Leu supply is high, the requirement per
se of Val is not affected by the Leu content but the response to
Val is even more increased (Gloaguen et al., 2012, Figure 5). In
addition, piglets avoid (decrease) to eat BCAA imbalanced diet (i.e
Val deficient). Indeed, Gloaguen et al. (2013) shown that a piglet is
able to detect within 1 hour after meal the dietary Val deficiency
and consequently reduces its feed intake. The combination of the
decreased feed intake and feed efficiency results in a drastic
decrease of the ADG.
Figure 4 - Global pattern of the branched-chain amino acids

metabolism (AEL, 2013).
Figure 5 - Effect of SID Val: Lys on piglets performance with

(Gloaguen et al., 2012) or without (Barea et al., 2009)
large supply of Leucine.
Among the BCAA, Val is the limiting AA in the piglet diets

when no L-Valine is added. It is therefore crucial to control the
minimum levels of dietary Val. The use of 70% SID Val: Lys
allows to counter act any imbalance in BCAA supply and to
maximize feed intake, growth and feed efficiency of piglets.
ISOLEUCINE REQUIREMENT IN PIGLETS
Isoleucine is a potential limiting AA after Val and its dietary

level could determine the dietary protein level in a formula using
the complete range of available feed-use AA. It is therefore
necessary to consider this AA in the formulation and to assess a
safe requirement level. The literature is important concerning Ile
but controversial results were published on the requirement
estimates. However, the factor of variation has been described

and today robust recommendations on Ile levels can be
established for commercial diets.
Isoleucine belongs to the group of the branched-chain AA
together with Val and Leu, but also to the Large Neutral AA group
(LNAA) which includes BCAA, Trp, His, Phe and Tyr. This group of
AA shares a common saturable transporter at the blood brain barrier
level. An analysis of literature about the Ile requirement in piglets
must take into account these factors. Van Milgen et al. (2013) ran a
meta-analysis work on the response of growing pigs to Ile and
concluded that the requirement is influenced by the presence of
spray-dried blood cells (SDBC) in the diet. Indeed, in these products,
the AA pattern is very imbalanced (Figure 6), and their use increases
the Ile requirement level. Due to their poor Ile contents, SDBC have
been extensively used in Ile dose-response studies but the
requirement estimates used to be very variable and very high in
comparison to requirement estimates based on blood-free diets.
Figure 6 - Amino acids profile of SDBC vs soybean meal; as % of

crude protein (CP).
A review of the Ile dose-response in piglets is proposed in

Figure 7; only the trials in which blood-free diets were used were
selected. Van Milgen et al. (2013) proposed a minimum
requirement of 50% SID Ile: Lys when blood-free diets are used.
For practical purpose, we recommend the use of 53% SID Ile: Lys.
When using blood cells attention must be paid to the dietary level
of Ile with an increased requirement.
Figure 7 - SID Ile: Lys dose-responses in piglets, blood-free diets.

Effect on average daily gain (ADG).
LEUCINE REQUIREMENT IN PIGLETS
Leucine is a BCAA, generally in excess supply in current

piglet’s diets, particularly in corn-based diets. Leucine plays a key
role in the catabolism of the BCAA. A Leu excess together with Val
and/or Ile deficiencies is described in the literature to be
detrimental for piglet performance (Bulletin 35, Corrent et al 2010).
Therefore, any imbalance within BCAA dietary supply must be
avoided. However, a recommendation of a leucine maximum is not
meaningfull since the issue lies in the balance with the others
BCAA. The most efficient way is to formulate closer to the leucine
requirement and to control the minimum Val and Ile levels. It is
therefore necessary to assess the animal response to Leu to
determine to what extent Leu levels can be reduced.
Literature about Leu requirement in piglets is scarce but
recent trials have been performed in Europe and are reported in
Figure 8. For instance, Gloaguen et al. (2012) published a study
about the Leu requirement in piglets and concluded that 102% SID
Leu: Lys was necessary on average to maximize the piglet
performance. However, as shown in Table 2, a 10% deficiency
slightly affects the response, which is confirmed by recent trials
presented in Figure 8.
Table 2 - Results of Gloaguen et al. (2012). SID Leu: Lys

requirement estimates and effect of a 10% deficiency
on performance, based on a curvilinear-plateau model
Gain to
Criteria to maximize ADFI ADG
Feed
SID Leu: Lys requirement estimates (%) 102.4 101.9 97.2
-10% deficiency affects the response by -1.9% -3.3% -2.0%
Figure 8 - SID Leu: Lys dose-responses in piglets. Effect on

average daily gain (ADG).
In their work, Soumeh et al. (2015) studied the BCAA dose-

responses with the same protocols. Each dose-response was
modeled using a curvilinear-plateau model and compared to each
other to determine the most responsive AA (Figure 9). This work
has been done only on ADG since BCAA have a strong impact on
the feed intake and FCR when diets are BCAA-deficient. In these
studies, Soumeh et al. (2015) determined requirements for Val at
71, Ile at 52 and Leu at 94% of SID Lys. Considering a level of -
10% of the estimated requirement for the ADG, the response to
Val is the strongest (-6%), followed by Leu (-5%) and Ile (-3%). For
Val and Ile, the results are similar to literature (van Milgen et al.,
2013). However, the Leu requirement is estimated at a lower level
than the one proposed by Gloaguen et al. (2013) (94% vs
103% Leu: Lys DIS) but the response is higher in this study (-5%
vs -3%). This suggests a Leu requirement between 94 and 103%
SID Leu: Lys. Based on these data a minimum level of 100% SID
Leu: Lys is recommended for piglet diets.
Figure 9 - Effect of increasing level of Ile, Val and Leu on ADG of

piglets, expressed in percentage of average plateau
value in each trial and using curvilinear-plateau model
(Soumeh et al. 2015).
On the X-axis, value 100 is SID requirements in ratio to Lys: 52%

Ile, 71% Val et 94% Leu. Model for Ile <0.52 : ADG = 448-
4718*(0.52-Ile:Lys)2; Model for Val:Lys <0.71 : ADG = 410-
6424*(0.71-Val:Lys)2; Model for Leu:Lys < 0.94 : ADG = 314-
1831*(0.94 – Leu:Lys)2. Above requirement, the ADG is fixed
(plateau).
HISTIDINE REQUIREMENTS FOR PIGLETS
Histidine is considered as an IAA for piglets but its

requirement has not been extensively studied. The current dietary
levels in piglets’ diets are not limiting and His could be limiting
after the BCAA group depending on the structure of the formula
(i.e. feedstuffs AA contents). To be complete, an ideal AA profile
must account for His level. In order to express the His requirement
within the ideal protein concept, Gloaguen et al. (2012) and
Wessels et al. (2016) performed His dose-responses in piglets in
which Lys was sublimiting. The response to ADG is presented in
Figure 10, and an average requirement of 32% SID His: Lys has
been assessed.
Figure 10 - SID His: Lys dose-responses in piglets, effect on

average daily gain (ADG).
Two other studies had been performed in the past (Table 3)

but the requirement estimates are expressed in different units
which makes difficult the synthesis. However, considering the
response when His is deficient, the estimate of 32% SID His: Lys
is safe to be used in low protein diets.
Table 3 - Literature available about the His requirement in piglets
Body Response Requirement

weight criteria estimates
Izquierdo et al., 1988 10-20 kg ADFI, ADG, G:F 0.36% total His
Li et al., 2002 10-20 kg ADFI, ADG, G:F 30% AID His:Lys
Gloaguen et al., 2012 10-22 kg ADFI, ADG, G:F 32% SID His:Lys
Wessels et al., 2016 8-20 kg ADFI, ADG, G:F 28 -32% SID His:Lys
PHENYLALANINE AND TYROSINE
Phenylalanine and Tyrosine are aromatic AA also classified

as IAA and semi-dispensable AA, respectively. The current piglet
diets still provide large amount of Phe and Tyr, even after the
dietary crude protein reduction which is currently achievable using
the available feed-use AA. The requirement of these AA had
therefore not been studied. Pioneer works are presented here.
Phenylalanine can be converted to Tyr by irreversible
hydroxylation through an enzyme which is activated by Phe itself.
Tyrosine is considered as a semi-dispensable AA because a Tyr
deficiency could occur when the dietary Phe supply is insufficient
to supply Tyr. Because Phe can be used for Tyr synthesis, the
requirement for the latter had not been explicitly quantified and the
requirement had been usually expressed in recommended AA
profile as the sum of Phe + Tyr. Recommended requirements
vary from 54 to 57% SID Phe: Lys and 93 to 111% SID
(Phe+Tyr):Lys. Gloaguen et al. (2014) worked on dose-responses
to Phe and Tyr and on the possibility to substitute Tyr by Phe. It is
shown that the minimum SID Phe: Lys requirement is about 54%
and the maximum SID Tyr: Lys requirement is about 43% (Table 4).
Table 4 - Results of Gloaguen et al. (2014). SID Phe and Tyr to

Lys requirement estimates and effect of a 10%
deficiency on performance, based on a curvilinear-
plateau model
Criteria to maximize ADFI ADG Gain to Feed
SID Phe: Lys requirement estimates (%) 54.8 54.2 52.6

-10% Phe deficiency affects the response by -1.8 -3 -1.7
SID Tyr: Lys requirement estimates (%) 42.8 39.7 -
-10% Tyr deficiency affects the response by -0.6 -0.7 -
Nevertheless, it seems that considering the sum of Phe+Tyr

to assess the requirement is not relevant since the substitution of
Tyr by Phe was not successful in this work (data not shown here).
This could be linked to the particular usage (oxidation) of the Tyr
obtained from Phe in comparison to the dietary Tyr. More research
is needed on this issue, and for practical feeds we still recommend
a minimum of 95% SID (Phe+Tyr): Lys with minimum SID Phe:
Lys of 55%.
IDEAL AMINO ACIDS PROFILE
In growing animals, the concept of “ideal protein” or ideal

AA profile is a concept where AA pattern (defined as a percentage
of lysine) maximizes growth, nitrogen retention or another
response criterion of interest. In this profile, all indispensable
amino acids are equally limiting for performance, just covering the
requirements for all physiological functions. Lysine has traditionally
been used as a reference because it is the first limiting AA for
growth in pigs. It is frequently assumed that the ideal protein
profile does not change for a given growing stage. In practical

nutrition, this offers the advantage that the lysine requirement will
vary (per kg of feed or per MJ of energy), but not the ideal amino
acids profile expressed relative to lysine. Each of these ratios can
thus be directly introduced as a constraint in feed formulation.
Table 5 - Ajinomoto Eurolysine s.a.s. (AEL) Amino Acids profile for

piglets and comparison to other recommended profiles
SID BSAS NRC VSP INRA Rostagno AEL

Values (2003) (2012) (2016) (2013)* (2011) (2013)
UK USA DK FR BR Europe
Lys:Lys 100 100 100 100 100 100

Thr:Lys 65 59 61 65 63 65
Met:Lys 30 29 32 30 28 30
(Met+Cys):Lys 59 55 54 60 56 60
Trp:Lys 19 16 20-22 22 18 22
Val:Lys 70 63 67 70 69 70
Ile:Lys 58 51 53 52** 55 53**
Leu:Lys 100 100 100 101 100 100
His:Lys 34 34 32 31 33 32
Phe:Lys 57 58 54 54 50 55
(Phe+Tyr):Lys 100 93 100 - 100 95
Tyr:Lys - - - 40 - -
Indicatives recommended SID Lys levels: From 1.35 to 1.15% for 6-12 and
12-25 kg piglet live weight respectively.
*Gloaguen et al. (2013)
**Requirement is given for blood-free diet
A comparison of the different recent recommended AA

profiles (Table 5) shows that there is still variability between the
different institutes. This is due for instance to different
methodological approaches. As an example, an important

difference between NRC (2012) and INRA (2013) can be
explained by the factorial method used by NRC and the empirical
approach used by INRA.
Others factors that lead to recommend an AA profile are
listed in Figure 11. Apart from the methodological aspects, the
difference between recommended AA profiles lies in the global risk
management of the feed formulation. In a context of general usage
of low protein diets as it is in Europe, the risk is taken on each AA
instead of crude protein, which is more relevant since crude
protein is not a predictor of the piglet’s performance.
Figure 11 - Example of factors to consider to assess an amino

acids profile for piglet commercial diets
LOW CRUDE PROTEIN DIETS
Knowing the amino acid contents of feedstuffs, their

digestibility, using net energy values, and implementing updated
ideal AA profile, allow to implement efficiently low protein diets in
piglets. The use of low dietary CP levels in piglets’ diets is
increasing in Europe for health, economic and environmental
reasons. This technic is already known and furthermore
implemented since the range of available feed-use AA has
increased (i.e. L-Valine). There is stillroom for further dietary CP
reduction in practical feeds without compromising growth
performance.
Using low protein diets in piglets: Efficient and safe
 A lower supply of dietary CP reduces the global amount of

undigested protein supplied in the distal intestine of piglets
and results in a better acidification (and digestion) of the
bolus. This ends by a lower proliferation of the pathogenic
bacteria and significantly reduces the occurrence of
diarrhea (Lordelo et al., 2008).
 By formulating on each IAA instead of minimum dietary CP,
the most balanced AA profile is supplied to the animal
avoiding lack and excess of AA, the amount of soybean
meal is reduced in the formula and substantive cost savings
are achievable. By applying a precise AA profile, risk
management can be implemented and the best
opportunities of using local feedstuffs and co-products can
be taken.
 Reducing dietary CP is one of the best available technics to
reduce nitrogen output into the environment: -1 point of
dietary CP results in -10% of nitrogen excretion.
Using the ideal AA profile recommended by Ajinomoto

Eurolysine s.a.s. and the available feed-use AA, dietary CP can be
reduced safely and efficiently as shown in the following trials

presented (Table 6).
Table 6 - Effect of lowering dietary crude protein (CP) on piglet

performance (trials using only available feed-use AA)
Body Dietary SID ADFI ADG FCR P Ideal AA

weight CP Lys (g/d) (g/d) profile
(kg) (%) (%) INRA 2013
Lordelo et al. (2008) 7-23 20.5 1.12 932 583 1.60 ns

17.0 1.12 941 571 1.65
Jansman et al. (2008) 10-25 19.0 1.03 869 567 1.54 ns
16.0 1.03 866 568 1.52 
Norgaard and 9-21 19.0 1.10 627 445 1.40 ns

Fernandez (2009) 17.0 1.10 635 449 1.41 
Vinyeta et al. (2010) 8-25 17.5 0.94 857 564 1.52 ns
15.5 0.94 873 567 1.54 
Gloaguen et al. (2013) 12-23 17.6 1.00 766 450 1.70 ns

15.6 1.00 775 454 1.71 
Jansman et al. (2013) 8-25 16.8 1.00 835 548 1.49 ns 
15.4 1.00 816 564 1.48
Rondia et al. (2016) 8-16 20.0 1.10 483 302 1.59 ns 
17.5 1.10 507 333 1.52
AEL Trial (2016) 8-20 17.0 1.05 826 452 1.80 ns 
15.5 1.05 800 442 1.81
A PIONEER WORK: FORMULATING DIETS FOR PIGLETS

WITHOUT SOYBEAN MEAL
After a complete review and work on AA requirements,

Gloaguen et al. (2013) proposed to test to what extent the CP content
of piglets diets can be reduced by substituting soybean meal by
wheat, barley, corn and free AA, without affecting performance. The
study is based on the Ajinomoto Eurolysine’s AA profile. In the first 4
diets, soybean meal was still used and CP decreased from 17.6 to
11.8%. In additional 2 diets, only cereals and free AA were used to
test the effect of nitrogen addition (13 vs 14% CP by L-Glu addition in
treatment 6). The formulas and results are summarized in Table 8.
Table 7 - Effect of reducing dietary crude protein content on

performance of piglets (Gloaguen et al., 2013).
Treatments 1 2 3 4 5 6
Feedstuffs used CEREALS - SBM - L-AA CEREALS & L-AA
Cereals (%) 50 60 80 85 80 80
Soybean Meal (%) 25 20 10 2 0 0
L-Lys HCL (%) 0.28 0.46 0.70 0.92 1.00 1.00
DL-Met, L-Thr, L-Trp + ++ +++ ++++ ++++ ++++
L-Val - + ++ +++ ++++ +++
Others L-EAA - - + ++ +++ +++
Others L-NEAA - - - - + ++
SID Lys (%) 1.00 1.00 1.00 1.00 1.00 1.00
Ideal AA profile      
Crude Protein (%) 17.6 15.6 13.5 11.8 13.0 14.0 P
BWi, kg 12.7 12.7 12.6 12.6 13 12.8 0.60

BWf, kg 22.2a 22.2a 21.9a 20.1b 21.8a 22.3a <0.01
ADFI, g/d 766 775 779 734 810 782 0.55
ADG, g/d 450a 454a 442a 358b 450a 451a <0.01
G:F, g/g 0.59a 0.59a 0.57a 0.49b 0.52b 0.58a <0.01
 The feed intake is not influenced whatever is the protein

level,
 Growth performance are maintained till the level of 13.5 –
14.0% CP (treatments 1, 2, 3 & 6)
 Comparing diets 5 and 6 shows that at 13% CP, nitrogen
was limiting: Adding a N source up to 14% CP allows to
recover performance as in treatment 1,
 This trial indicates that efficiency of utilizing AA is not lower
for free AA compared with protein bound AA in pigs offered
feed ad-libitum,
 In the successful treatment 6, 78% of the dietary SID Lys
content was supplied by L-Lys-HCL.
This trial shows that there might be a minimum of N

between 13 and 14% CP (for 1.00% SID Lys). However, it
confirms that tremendous dietary protein reduction is achievable in
current piglet feeds. It indicates also that when using the
Ajinomoto Eurolysine ideal AA profile, dietary CP can be reduced
efficiently in piglet diets by using feed-use AA.
CONCLUSION
Indispensable amino acids, as essential nutrients, must be

supplied in the diets for piglets. As they predict performance, they
represent factors of variability of the growth and must be
monitored with care to get the best performance in variable
conditions. As a first step, amino acids contents in feedstuffs have
to be estimated and digestible values used as predictors. The
increasing knowledge about amino acid nutrition allows to
recommend a robust ideal amino acid profile that supports feed
intake, growth, nutrients efficiency and health challenges of the
piglets. Although an ideal AA profile gives fixed values, response
to an AA can vary depending on the conditions and more studies
are needed to refine the impact of external factors on an AA effect.
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AMINO ACIDS ESSENTIALITY FOR SOWS: NEW
APPROACHES
1 2
Márvio Lobão Teixeira de Abreu , Alysson Saraiva
1
Departamento de Zootecnia, UFLA. Lavras-MG. CEP: 37200-000;
2
Departamento de Zootecnia, UFV. Viçosa-MG. CEP:36570-000
SUMMARY
The high prolificacy of modern sows has allowed increasing

the number of pigs sold and the profitability of the production
system. However, greater metabolic burden of the sow and lack of
uniformity of the litters at birth and weaning has been observed. In
addition, it is recognized that such animals are more nutritionally
demanding.
The evolution of the nutritional concepts on amino acids
requirements of hyperprolific sows must acknowledge that some
amino acids, such as arginine, glutamine, glutamate, proline,
branched chain amino acids, among others, besides being
required for the synthesis of proteins (placenta, uterus, fetuses,
mammary gland, milk) may participate in important regulatory
pathways (secretion of hormones, increased vascularity, oxidative
status, immune response, etc.) that are most required during
pregnancy and lactation.
In this sense, we can conclude from the results of several
studies that such amino acids must have their concentrations
increased in the diets of pregnant and lactating sows to obtain the
maximum productive/reproductive performance. This will allow to
update nutritional requirements recommendations (tables/guides),
recognizing the essentiality of dietary amino acids, whether
essential or not, according to the classic definition of essential
amino acid.
INTRODUCTION
Moderns ows are more productive and are known as

hyperprolific. Therefore, they have increased
physiological/metabolic needs that may have implications on their
nutritional requirements, particularly amino acids. Besides
participating in the structural formation of proteins of the body,
amino acids can act as regulators of gene expression, immune
response, hormonal regulation, redox status balance, etc. The
better understanding of these other functions has attracted the
attention of nutritionists as these metabolic demands could be
translated into nutritional demands.
The evolution of the nutritional concepts related to amino
acids essentiality goes beyond the classic concepts of their
synthesis in adequate quantity or not in the body. In this way, we
have tried to evaluate the response of sows to the
supplementation of some amino acids recognized as functional.
This work discusses current aspects of
physiological/metabolic needs of pregnant and lactating sows and
their implications on the requirements of some amino acids.
Although these requirements are undefined, the supplementation
of some of them will enable to obtain maximum
productive/reproductive performance of the modern sows.
UNDERSTANDING THE METABOLIC DEMANDS OF

HYPERPROLIFIC SOWS AND THE PARTICIPATION OF
FUNCTIONAL AMINO ACIDS
Genetic and environmental aspects will not be addressed,

only physiological and metabolic factors that can influence
nutritional requirements. Besides the structural need (easy to
quantify), aspects of the physiological participation of some amino
acids that may justify their adjustments in the diets will be
presented.
Gestation
From the classical concept of structural function of amino

acids, during gestation the needs of these nutrients are high and
influenced by parity order (Young et al., 2005), gestational stage
with its different physiological phenomena (Kim et al., 2009), and
by the number of piglets present in the uterus (Wu et al., 2015).
Thus, all amino acids are required in large quantities in the diets to
meet the nutritional needs for growth of young pregnant females,
development of the mammary gland, and for growth of a large
number of fetuses in the final third of gestation.
The current scenario is of large litters at birth which has
caused a metabolic overload to the uterus and placenta of sows.
The importance of these organs was demonstrated by Freking et
al. (2007) as they selected sows for higher uterine capacity and
placental weights. Compared to females with higher ovulation
capacity, those females had higher number of fetuses per uterine
horn. According to Bazer et al. (2009), inadequate uterine capacity
is responsible for fetal deaths and/or suboptimal growth of fetuses
after 30 days of gestation.
The lack of piglets’ growth uniformity of sows supporting
large litters can be explained by lower uterus-placenta blood flow
per fetus (Pere & Etienne, 2009) and lower rate of amino acids
transport through placenta (Wu et al., 2006). This may be related
to sub-optimal nutrition of the sow, affecting the development of its
placenta, favoring the occurrence of small piglets at gestational
age or intrauterine growth retardation (IUGR).
During gestation, in addition to the amino acid requirements
for tissue accretion, some physiological functions related to the
uterus and placentas are requested in greater intensity. These
organs need to produce hormonal and growth factors that promote
embryo implantation and development, to synthesize antioxidant
factors for the maintenance of oxidative homeostasis in embryos
and fetuses, and also act efficiently in the transport of nutrients
and gas exchange. Some amino acids may actively participate in
these functions, such as arginine, glutamine, glutamate,

methionine, cysteine, etc.
Amino acids of the arginine family (arginine, proline, and
glutamine) are essential substrates for the proper development of
placenta, embryos, and fetuses (Wu et al., 2004; Liu et al., 2012),
since arginine is precursor of nitric oxide and polyamines. Nitric
oxide and polyamines (putrescine, spermine, and spermidine) are
essential for placental growth and angiogenesis (Wu et al., 2009),
increasing the availability of nutrients to the fetus. Polyamines,
which are synthesized in the porcine placenta from proline-derived
substrates, regulate DNA and protein synthesis, being directly
related to cell proliferation and differentiation (Wu et al., 2005).
Kong et al. (2014) reported that putrescine stimulates protein
synthesis in pig trophectoderm cells.
The placenta synthesizes and releases large amounts of
glutamine into the fetal circulation (Self et al., 2004). The absence
of glutaminase activity in the placenta of pigs contributes to this
(Wu et al., 2015). The importance of glutamine for fetal
metabolism was demonstrated by Wu et al. (2013). The authors
verified that glutamine is the amino acid in the circulation with
highest uptake by the uterus, but only 23% are retained in fetal
tissues. This indicates that glutamine is actively metabolized by
the fetus and can serve as regulator of metabolic pathways in
addition to being used for protein synthesis. Like arginine,
glutamine also activates the mTOR(mammalian target of
rapamycin) signaling pathway in the placenta cells (Kim et al.,
2013).
According to Wu et al. (2011) glutamine deficiency is the
main triggering factor for development of LPID and/or IUGR
piglets. In addition to energetic substrate for high proliferation cells
(Marc Rhoads & Wu et al., 2009), glutamine is a precursor of
nucleotides, amino sugars, also act as a signal for cell proliferation
(Yamauchi et al., 2002).
Glutamate is the major precursor of glutamine in the pig
placenta, establishing an important placenta/fetus cycle during
pregnancy (Wu et al., 2015). It should be noted that not all key
functions of glutamine can be met via glutamate (Wu 2009). Other
glutamine precursors are branched-chain amino acids (BCAA)
(Self et al, 2004).
Methionine plays an essential role during gestation as it
participates in the donation of methyl groups for DNA synthesis
and methylation (Locasale, 2013) during embryonic/fetal
development, which makes it extremely important for the
regulation of gene expression.
Due to the genetic selection for greater prolificacy,
metabolic loads increased significantly in modern sows, especially
at the end of gestation and during lactation, so that these females
may be in oxidative stress during these periods (Bernardi et al.,
2008; Berchieri-Ronchi et al., 2011; Zhao et al., 2013). In addition,
antioxidant nutrients decrease during pregnancy, resulting in a
significant fall at 110 days of gestation, beginning to normalize
only at the end of the lactation period (Berchieri-Ronchi et al.,
2011; Zhao, 2011).
Glutamate and glycine participate in the enzyme glutathione
peroxidase, an antioxidant peptide that exerts a beneficial function
in the control of lipid peroxidation and in the prevention of
deleterious effects of free radicals on the health of the sow and
fetuses (Salvolini et al, 2012). This may imply in increased needs
of glutamate and glycine supplementation in the diet.
In pigs, the reduced concentration of leucine in the umbilical
vein (Lin et al., 2012) and the decreased valine concentration in
the jejunum of the fetuses (He et al., 2011) are associated with
abnormal energetic metabolism in piglets with IUGR. In addition,
transport and fetal use of BCAA were significantly altered by
IUGR. This is important because, besides being involved in
muscle protein synthesis, through mTOR, the BCAA, as previously
seen, are precursors of glutamine in the pig placenta.
Lactation
During lactation, most amino acids are targeted to

mammary gland growth and milk production. The latter is
influenced by the feed intake capacity of the sow which can be
affected by thermal environment, handling, parity order, and
number of piglets disputing teats. It should be born in mind;
however, that the adequate supply of amino acids should allow
sows’ weight loss that do not compromise their growth (first parity)
or their future reproductive performance (first parity and
pluriparous sows).
The amino acids required for the synthesis of porcine milk
protein are derived from the bloodstream or synthesized from
precursors in the mammary epithelial cells (Manjarim et al, 2014).
According to Rezaei et al. (2016) these amino acids mustpermit a
synthesis of more than 0.600 kg of protein/day in a sow producing
13 kg of milk per day approximately.
The main proteins of sow milk are casein α-S1 (42.0%) and
β-lactoglobulin (20.0%), rich mainly in glutamate, glutamine,
proline, and leucine (Rezaei et al., 2016). This suggests high
uptake or intense metabolism of their precursor amino acids,
indicating the importance of their supply via circulation. For
example, high concentration of glutamate and glutamine can be
explained by the intense catabolism of BCAA in the mammary
gland of the sow (Li et al., 2009) and the high concentration of
proline is due to arginine catabolism in the cells of the mammary
gland. About 81%of arginine from arterial blood is catabolized in
the mammary gland (Li et al., 2009).
Glutamate, glutamine, and proline are used by the newborn
piglet enterocytes with different but important purposes for
maintenance of intestinal function, contributing to the growth of
suckling piglet. In fact, glutamine promotes intestinal growth and
muscle protein synthesis in neonatal piglets (Wang et al., 2008; Xi
et al., 2011). In turn, proline is used for the synthesis of citrulline,
which is converted to arginine, either locally or in the kidneys (Wu
et al., 2009). This partially alleviates the arginine deficiency of the

sow’s milk.
Amino acid metabolism plays an essential role in milk
production by regulating maternal endocrine status, redox status,
blood flow rate to the mammary gland, and by activation of the
Mtor signaling pathway (Wu, 2013).
Arginine can act as a secretagogue stimulating the
production of insulin, prolactin, and growth hormone which are
necessary for mammary gland development (Reyes et al., 1994).
This action was demonstrated by Mateo et al. (2008) by
supplementing arginine in the diet of lactating primiparoussows.
The authors found increased plasma concentrations of insulin and
suggested that according to them this hormone stimulates the
synthesis of nitric oxide, which in turn increased the blood flow of
the mammary gland, providing more nutrients for synthesis of milk.
In fact, the total milk protein in the study of Mateo et al. (2008) was
increased and a similar result was obtained by Moreira (2014). In
both studies it was verified an increase in the litter weight gain as
a result of arginine supplementation in the diet.
According to Manjarin et al. (2014) the understanding of the
various metabolic pathways of the mammary gland involving
amino acids must allow the update and definition of the amino acid
requirements of hyperprolific lactating sows.
AMINO ACIDS SYNTHESIZED IN THE BODY DIETETICALLY

ESSENTIAL FOR SOWS
The dietary essentiality of nonessential amino acids was

demonstrated by Deng et al. (2009) with piglets in the post-
weaning period (Table 1). According to the authors,
supplementation of essential amino acids in diets with high crude
protein reduction did not restore the performance of the pigs.
Table1 - Non-essential amino acid deficiency and post-weaning

piglet performance
Crude protein, %
Item
20.7 12.7 + Essencial aas.
Protein synthesis, %/day
L. dorssi 11.8 7.1
Liver 83.5 63.0
Pancreas 76.4 62.7
Kidney 36.1 24.1
Feed intake, g/day 432 455
Weight gay, g/day 299 264
Feed conversion 1.44 1.72
Thus, it is possible to conclude that amino acids, even

those synthesizable in the animal body, if present in limiting
amounts impair animal growth and, therefore, must be
supplemented in the diet, making them dietetically essential. The
same can happen with gestating and lactating sows with high
metabolic demands. Such demands may require the participation
of specific amino acids. In this way, the response of the sows to
the supplementation of some known functional amino acids has
been evaluated.
Gestation
In gestations considered as normal, most amino acids have

a positive relationship between their concentrations in maternal
plasma and fetal plasma, suggesting it is possible to increase their
availability to fetuses by raising their concentrations in the
circulation (Lin et al. (2014). In this sense, dietary amino acid
supplementation may be a useful tool in the nutrition of pregnant
sows.
Table2 - Positive results of the use of arginine and/or glutamine in

diets of pregnant gilts
Item Amino Amino Improvement,

Control Study
acid acid %
Bérard and
Number Arginine 9.30 13.00 29.00
Bee, (2010)
of viable fetuses
Arginine 10.5 12.40 18.00 Li et al., (2013)
Total weight of Bérard and
Arginine 3.73 4.93 24.00
viable fetuses, kg Bee, (2010)
Coef. of variation of
Quesnelet
the weight of piglets Arginine 25.40 20.60 19.00
al.(2014)
born alive
Mateo et al.
Arginine 1.86 0.66 35.00
(2007)
Stillborn piglets Arginine +
1.13 0.57 50.00 Wu et al. (2010)
Glutamine
Arginina 1.44 0.50 66.00 Liu et al. (2012)
Arginine +
11.03 11.90 8.00 Wu et al. (2010)
Glutamine
Total born piglets Arginine + Gao et al.
12.46 13.77 10.00
Glutamine (2012)
Arginine +
10.63 12.44 17.00 Li et al. (2015)
Glutamine
Arginine + Wu et al.,
9.91 11.33 13.00
Glutamine (2010)
Gao et al.
Piglet born alive, kg Arginine 11.25 12.35 9,00
(2012)
Che et al.
Arginine 10,19 11,81 16,00
(2013)
Mateo et al.
Arginine 13.19 16.38 24.00
(2007)
Arginine +
14.60 16.00 9.00 Wu et al. (2010)
Glutamine
Litter weight at Glutamina 13.70 14.70 7.00 Wu et al. (2011)
birth, kg Arginine 14.12 16.26 15.00 Liu et al. (2011)
Gao et al.
Arginine 15.82 17.52 10.00
(2012)
Che et al.
Arginine 16.03 18.10 13.00
(2013)
Controlling feed delivery during pregnancy in order to avoid

overweight of the sows may provide insufficient provision of some
amino acids for the fetuses. A number of studies have indicated
the supplementation of specific amino acids, mainly arginine and

glutamine, in the gestation diet has allowed improving the
productive/reproductive performance (Table 2).
The benefits obtained with arginine supplementation in
gestation diets have the following characteristics: a) the increase
in the number of piglets born is more significant in less prolific
sows; b) in these sows, the increase in litter weight at birth is
mainly due to the increase in the number of piglets born; c) in
hyperprolific sows the impact on piglet weight at birth is lower,
standing out the reduction of stillborn piglets and greater litter
uniformity birth, as a result of the reduction of low-birth weight
piglets.
The occurrence of low-birth weight piglets (less than 800 g)
is the main challenge in today's production systems. Low birth
weight and high weight variability of these piglets are associated
with lower neonatal survival, greater disease susceptibility,
reduced postnatal growth rate, and poor carcass quality (Wu et al.,
2006). This implies in extra handling costs and efforts in the pig
farm.
Glutamine is also effective in increasing prenatal survival
and reducing the occurrence of low-birth weight and/or IUGR
piglets. Its association with arginine needs further studies as no
additional effects have been observed.
To date, little is known about the dietary requirement of
non-essential amino acids by animals (Wu et al., 2014). However,
the dietary essentiality of some of these amino acids, that is, the
need of their supplementation in gestation diets was proposed by
Wu et al. (2013) from the recommendations of some studies
(Table 3).
Table3 - Recommendations of some non-essential amino acids

requirements during pregnancy
% of diet
Item Studies
<90 days of gestation >90 days of gestation
Arginine 1.13 1.53 Mateo et al.
Glutamine 1.22 2.22 (2007);
Glutamate 1.07 1.57
Proline 1.03 1.53 Wu et al. (2010)
According to Wu et al. (2014) one must consider

supplementing arginine throughout gestation and the other amino
acids listed in Table 3 in the final stage of gestation.
Studies with BCAA during pregnancy have involved other
animal models. Leucine-rich diets have recently been used to
minimize reductions in placental and fetal growth and to improve
muscle protein synthesis in rats (Viana& Gomes-Marcondes,
2013). Zheng et al (2009) also demonstrated that prenatal and
postnatal supplementation with BCAA stimulates muscle
development in neonates with IUGR.
Lactation
Studies with non-essential amino acids and their effects on

performance of sows and their litters during lactation are fewer in
comparison to those of pregnant females. But the goals are the
same, to explore the structural regulatory functions of those amino
acids, in order to increase the functionality of the mammary gland.
These functions may result in increased production or
improvement in the nutritional quality of milk. Likewise, the most
studied amino acids are arginine and glutamine.
In studies by Wu et al. (2004), it has been shown that milk
is deficient in some amino acids such as arginine, which may
compromise the development of piglets. According to estimates by
Wu et al. (2004), the sow’s milk supplies less than 40% of the total
arginine required daily by piglets at day seven of lactation.
Although studies with arginine supplementation in lactation

diets do not prove its increase in milk, favorable results have been
achieved. Mateo et al. (2008) verified an improvement in the
growth of suckling piglets from sows that had access to a diet
supplemented with 1.0% L-Arginine. According to the authors the
positive results were due to improvement son the amino acid
profile of sows’ milk. Likewise, Moreira (2014) associated the
greater weight gain of the litter to the improvement of the protein
and energy profile of sows’ milk.
The milk of the sow is rich in glutamine and this amino acid
is intensely used by the intestine of suckling piglets. Besides
important energy source for enterocytes, glutamine is a precursor
for the synthesis of purines and pyrimidines that act on cell
proliferation (Wu & Knabe, 1994). It is possible, according to
Manso et al. (2012) to increase the concentration of glutamine in
the milk of sows, favoring the growth of the piglets.
As muscle is an important supplier of glutamine to the
mammary gland, it may be depleted in certain situations, which will
result in increased sows weight loss, and impairment of future
reproductive performance. In fact, Lobley et al. (2001) found a
decrease of about 25% glutamine in the muscle of lactating sows.
Lactating gilts supplemented with L-glutamine had lower lean body
loss and higher concentration of glutamine in milk (Manso et al.,
2012). This increase can be accompanied by an increase in the fat
content of milk, from colostrum to the end of lactation (Santos de
Aquino, 2014).
Monosodium glutamate (MSG) was studied in diets of
lactating sows (7 to 28 days) by Rezaei et al. (2013). The authors
found that supplementation with 2.0% GMS resulted in lower
mortality rates of suckling piglets and heavier piglets at weaning.
In Table 4 are presented some studies with positive results
on the use of arginine and/or glutamine in diets of lactating sows.
Table4 - Positive results of the use of arginine and/or glutamine in

diets of lactating sows
Improvement,
Item Control Aminoacid Estudy
%
Pérez
Sow body weight loss, Arginine 16.28 7.48 54.00 Laspiur&Trottier
kga (2001)
Arginine 5.26 5.66 7.60 Mateoet al. (2008)
Weight of piglets at Glutamine 5.40 5.77 6.80 Wu et al. (2011)

weaning, kg Arginine 6.30 6.60 4.80 Moreira (2014)
Arginine 0.183 0.202 10.40 Mateoet al.(2008)

Daily weight gain of
Arginine 0.166 0.185 11.44 Lima (2010)b
piglets, kg
Arginine 0.250 0.263 5.20 Moreira (2014)
Number of weaned
Arginine 10.46 11.52 10.13 Lima (2010)b
piglets
a b
Hot environment; First week of lactation.
CONCLUSIONS
The hyperprolificacy of the modern sows has led to greater

efforts in the nutritional planning of the females stock in pig farms.
The guideline should be toadequate the diet to the nutritional
requirements of the sows.
The evolution of the nutritional concepts related to the
amino acid requirement of hyperprolific sows must allow to
recognize that besides being necessary for maintenance and
synthesis of tissues, some amino acids are required for production
of important metabolites for certain physiological functions in
addition to having regulatory functions in the animal body. This
implies deepening the knowledge about the need for some amino
acids important for such metabolic demands and whether it can be
translated into nutritional requirements, making these amino acids
dietary essential.
From several studies we can conclude that some amino
acids, such as arginine, glutamine, glutamate, branched-chain
amino acids, among others, must have their concentrations in the

diets of pregnant and lactating sows increased when it is desired
to obtain maximum productive/reproductive performance.
The feeding period and quantity provided should be clearly
defined from the desired benefit to achieve to increase specific
productivity indicators in different pig production systems.
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AMINO ACIDS: A TOOL TO IMPROVE MEAT
QUALITY
Pierre-André Geraert
Dolores I. Batonon-Alavo & Yves Mercier
Adisseo France S.A.S., 92160 Antony, France
ABSTRACT
Amino acids have largely been shown to be the key nutrients

for muscle growth and thus meat production. The tremendous efforts
of geneticists have greatly enhanced the muscle growth in pigs and
poultry and the efficiency and sustainability of animal protein
production. However, consumer demand is evolving toward better
food, and thus quality has become increasingly important. Until
recently, the amino acid profile of the muscle was largely considered
to be determined by genetics and with the rather poor influence of
dietary composition. Recent experiments show the influence of amino
acid supply, even short-term exposures to single amino acids, on the
protein composition and meat quality parameters of pork and broiler
muscles. Such an approach will allow animal protein producers to take
into account nutritional progresses and better value their products.
Moreover, improving meat quality also means addressing better meat
preservation and maintenance of quality parameters through the retail
chain. Recent works show the benefits of dietary amino acids on meat
preservation to oxidation, for instance, guaranteeing better meat
stability and keeping its organoleptic properties at their optimal figures.
However, high growth rates and enhanced breast meat yields have
seen the emergence of new meat defaults: White Stripping and
Wooden Breast. Their etiology is under scrutiny but is not yet fully
understood. These muscular pathologies or defaults emerge during
the growth of the chickens and might involve the amino acid supply
and redox balance. This is a key issue to solve to maintain consumers’
interest in broiler meat and its high nutritional and hygienic quality.
INTRODUCTION
The growth of the world’s human population is increasing

the demand for animal protein worldwide. Meat production has
continuously increased through genetic selection and progress in
health, farm-management practices and nutrition. Supplementing
farm animals’ diets with amino acids is a current practice to
optimize growth and the feed conversion ratio and improve the
sustainability of animal protein production through the reduction of
dietary crude protein levels and, consequently, the nitrogen
excreted. Most often, the objective of dietary amino acid
supplementation is only to optimize production performance.
However, consumer desires are constantly evolving, and once the
first step of affordable meat is obtained, the request is to obtain a
good-quality meat. Depending on countries and consumers habits,
meat demand has progressively evolved from “fresh products”
(ready-to-cook) to “more elaborated products” (ready-to-eat).
These changes greatly influence breeding methods, which are
increasingly taking into account the quality aspects of the animal
products to satisfy consumers’ demands.
Since the 60s, meat science has long focused on managing
perimortem events such as transport, slaughter conditions,
refrigeration conditions, packaging and so on. More recently,
scientists have demonstrated links between breeding and/or
dietary treatments with meat quality parameters and their evolution
during storage. Studies on relationships between dietary amino
acids and meat quality are even more recent and began in the
early 2000s. The present review will address the different aspects
of meat quality traits in relation to dietary amino acids and the
possibilities to use amino acid supplementation to improve meat
quality.
MEAT QUALITY TRAITS AND CONSUMER DEMANDS
Meat quality may have different meanings depending on the

retailer’s final objective. According to consumers’ habits and
regional usage, broiler meat can be proposed as whole broiler, cut
parts or cooked and more elaborate products that require different
quality criteria. The same difference also exists for pork with even
more meat transformation through the delicatessen “charcuterie”
processing that has been developed in some countries. In
chickens, although difference depending on culture, the focus on
white meat (breast, fillets, and loins) has pushed the animal
industry to largely increase its production. Beyond hygienic
aspects and nutritive value, meat-quality traits are based (i) on
visual aspects that consumers can assess at retail places such as
color and drip loss and (ii) on organoleptic aspects such as
tenderness, juiciness, odor and flavor. Moreover, depending on
the retail form of the meat and packaging process (vacuum,
modified atmosphere packaging or simple retail film), visual and
organoleptic properties can be largely modified (Renerre, 1990).
Meat color and its stability during the retail period is
certainly the most impactful factor for consumers’ buying decision.
Meat color is dependent on muscle pigment, mainly myoglobin
with its content and its reduction/oxidation (redox) status. Color
criteria have been extensively studied in beef meat due to its high
quantity of myoglobin and the rapid change of redox status that
changes the color from a bright red (highly appreciated) to a
brownish color that consumers systematically reject. As color
stability problems are mainly linked to myoglobin content, they are
thus linked to muscle type and species that present different
myoglobin levels (Surendranath & Poulson, 2013). Surface
spectrometers allow determination of the three components of
meat color, i.e., L*, a* and b*, respectively, for lightness, redness
and yellowness, and its evolution during storage.
Drip loss and, consequently, the amount of exudate in the

retailing package can also impact consumers’ buying decision. If
color stability is certainly the first aspect for beef meat, drip loss is
a bigger problem in pork and poultry meat (Pearce et al. 2011).
The assessment of this quality criteria is easy to measure as the
difference of the weight of a piece of meat after a period of time.
This parameter might also have other applications, as it is well
correlated with water holding capacity (WHC), which is an
indicator of cooking losses and is particularly important in the case
of further processing.
Among the organoleptic aspects, tenderness is an
important factor for the determination of meat quality. This factor is
linked to muscle type and maturation time, allowing the
tenderization process to occur. This quality trait is more relevant
for beef and pork meat but is not an issue for poultry meat or fish
(Ouali et al. 2006). Tenderness is under the influence of the
collagen amount in the muscle that will determine the optimal
cooking process: grilling when the collagen content is low or
boiling when the collagen content is high. However, tenderness is
also dependent on the maturation process, which involves
intramuscular proteolytic systems (e.g., cathepsin, calpain,
proteasome), which degrade myofibrillar structures during
refrigerated storage and allow decreases in the toughness of the
meat.
Finally, organoleptic aspects such as odor and flavor
depend on the lipid content of the muscle and oxidative status.
This latter aspect will be more discussed in the oxidative process
section. Interestingly, of these four quality aspects of the meat, at
least three depend on the ultimate pH of the meat and are highly
interdependent.
FROM MUSCLE TO MEAT
What is the limit between muscle and meat? When does

muscle turn into meat? Slaughter is obviously the first step of the
process of muscle transformation. After slaughter and
exsanguination, peripheral tissues such as muscles are deprived
of the nutrients and oxygen that trigger this process. After death,
muscle metabolism will be turned in an anaerobic condition that
leads to lactic acid formation as the end product of glycolysis
(England et al., 2013). This lactic acid accumulation, due to the
lack of evacuation of the products by blood flow, results in a pH
drop in the muscle. This pH drop is dependent on the muscle’s
glycogen reserves, which are affected by many factors like feed
withdrawal, muscular activity before slaughter and genetic traits.
Under “standard” conditions (if “standard” can be applied to the
diversity of practices), the pH drop of the muscle starts a few
minutes after the death of the animal until an end point, which is
generally assessed 24 hours after slaughter and named “ultimate
pH” (pHu). For broiler breast fillets, pHu varies between 6.1 and
5.7. Above 6.1, the meat is considered dark firm dry (DFD),
whereas below 5.7, the meat is considered pale soft and exudative
(PSE). Texture, color and drip loss (or exudation) are linked to
pHu. Indeed, a low pH is associated with high lightness (L) values,
high exudation levels and softness. These kinds of meat are also
described for low transformation rates during the cooking process
associated with low WHC (Berri et al., 2015), whereas high pH
values are associated with firm texture, low L values and low
exudation. Moreover, besides the organoleptic problem of a high
pH, higher microbial proliferation and poor conservation are
observed. Another consequence of the pH drop and energy
depletion of muscle tissue is muscle shortening, resulting in
myofibrillar contraction. During this process, some extracellular
water will also be expulsed from the tissue, thus constituting the
first weight loss of the muscle. This muscle shortening, also called
rigor mortis, leads to the tougher state of the muscle that will be
attenuated during refrigerated storage and the maturation process.

During this process of maturation, the endogenous proteolytic
systems will breakdown myofibrillar proteins that will decrease
hardness (Ouali et al., 2006). During this process, almost all of the
proteolytic systems are involved, including cathepsins, calpains,
proteasomes and caspases. This process’ duration is largely
different between species and lasts for few hours in poultry up to a
few weeks for beef. Moreover, depending on the initial lipid
content of the muscle, the enzymatic activity and level of oxidation
flavor develop during this phase. The complexity and interactive
aspects of meat-quality traits are presented in Figure 1 (Geay et
al., 2002).
Figure 1 - Factors affecting meat quality (according to Geay et al.,

2002).
Amino acids: Key for protein deposition
Amino acids are mainly released from dietary protein

digestion, absorbed from the gastrointestinal tract, and
metabolized to support both anabolic and catabolic reactions.
Amino acids are building blocks for tissue proteins and essential
substrates for the synthesis of many other substances of
physiological importance (e.g., glutathione, creatine, carnitine,

thyroid hormones) (Kong et al., 2012). The catabolism of amino
acids will produce nitrogen eliminated as urea in mammals or uric
acid in poultry (Figure2). The continuous synthesis and
degradation of proteins is termed protein turnover, which
determines protein balance in tissues.
Figure 2 - Metabolism of amino acids (adapted from Larbier and

Leclercq, 1992).
Maximising the protein deposition in growing pigs and

broilers is nutritionists’ main objective. Although protein
metabolism is regulated by numerous factors such as
physiological state, genetic and environmental factors and
hormones, amino acids are recognized to be essential in such
regulation. The major determinant of protein deposition is the
dietary supply of amino acids. Due to its large mass, skeletal
muscle is the largest reservoir of both protein-bound and free
amino acids in the body (Wu, 2009). Several authors have

examined muscle development as a response to incremental
amino acids doses or crude protein-level variation. Tesseraud et
al. (2003) showed muscle weight response under low (12% CP)
and moderate (21.6% CP) dietary protein-supply conditions in
broilers receiving adequate energy levels. The reduction of crude
protein was accompanied by a reduction of amino acid levels in
the diet. Birds given the high-protein diet grew faster, ate less feed
and had a better feed utilization than their counterparts given the
low-protein diet. A significant reduction of circulating
concentrations of insulin-like growth factor-I (IGF-I) was observed
with low dietary protein. IGF-I is known to stimulate differentiation
and proliferation of satellite cells, thus contributing to lean tissue
growth (Doumit et al., 1993). Consequently, the restriction of
dietary protein reduced the pectoralis major (breast muscle) and
sartorius (leg muscle) (Tesseraud et al., 2003). Similarly, protein
synthesis is affected when an insufficient level of a specific amino
acid is provided. Diets with low lysine (Lys) contents were found to
limit breast meat formation by reducing protein accretion from
protein synthesis in broilers (Tesseraud et al., 1996; Costa et al.,
2001). A more pronounced decrease of protein accretion was
observed with methionine (Met) and cysteine (Cys) deficiency in
comparison to the deficiency of Lys, histidine or tryptophan in
broilers (Kino et al., 1986). In pigs, as Conde-Aguilera et al. (2015)
demonstrated, protein mass and synthesis are reduced when Met
and Cys are deficient in the diet. Accordingly, Chang and Wei
(2005) found that lysine deficiency in post-weaned piglets reduced
fractional protein synthesis in different muscles. Protein synthesis
can occur only when all the amino acids needed are present
simultaneously. Thus, for example, the supplementation of Met in
a deficient diet resulted in an increase of protein synthesis and
accretion in the muscles, e.g., for the gastrocnemius and
pectoralis major muscles in chicks (Barnes et al., 1995).
Moreover, signalling pathways that regulate protein
metabolism have been intensively studied. One of the most
explored is the mammalian target of rapamycin (mTOR) pathway,

and strong evidence exists that amino acids act as regulators of
metabolic pathways. Leucine has been found to act as a signalling
molecule to stimulate protein synthesis in skeletal muscles and
may reduce protein degradation. The underlying mechanism is the
activation of the mammalian target of the rapamycin complex 1
(mTORC1) signalling pathway (Anthony et al., 2001; Suryawan et
al., 2011; Kao et al., 2016). Conversely, the other two branched-
chain amino acids, valine and isoleucine, have no impact on
mTOR phosphorylation. Among those amino acids with signalling
function, Met was shown to induce p70S6K activation in mammals
(Stubbs et al., 2002). A study by Stubbs et al. (2002) on ovine
hepatocytes demonstrated that Met supply affects growth-
hormone-induced IGF-I gene expression. In addition, Met plays a
specific role in mRNA translation and is a source of methyl groups
that are used for DNA methylation. A deficiency in Met may inhibit
the first initiation of the mRNA translation step and affect protein
synthesis. Consequently, the supplementation of adequate levels
of amino acids is important to ensure that pigs and broilers
express their potential of meat production.
AMINO-ACID DEFICIENCY AND MUSCLE COMPOSITION
Amino acids are levers to improve growth performance and

muscle growth. In broilers, Met and Cys are the first limiting amino
acids in practical diets, whereas in pigs, they are considered the
third limiting amino acid for cereal-soybean-based diets. Many
studies show that Met supplementation, either as DL-Met or OH-
Met (HMTBA), to a deficient basal diet improves broiler-growth
performance (Dozier and Mercier, 2013; Agostini et al., 2015) and
carcass yields (Fatufe and Rodehutscord, 2005). In pigs, studies
have focused on lysine (Apple et al., 2004; Auldist et al., 1997),
threonine (Ettle et al., 2004) and tryptophan’s (Capozzalo et al.,
2012) effects on performance and carcass characteristics. More
recently, the supplementation of branched-chain amino acids into
low-protein diets has gained interest (Barea et al., 2009; Craig et

al., 2016). Though it is clear to nutritionists that all amino acids are
required for animals’ growth, there is still a need to clarify the
consequences of moderate deficiency. In other words, what will
happen if the third or the fourth limiting amino acid is not supplied?
Animals have requirements for amino acids classified into
essential and nonessential amino acids. Essential amino acids
must be supplied through nutrition, whereas animals can
synthesize the latter group de novo. These requirements are
usually expressed as ratio to lysine (Baker, 1997), which is
supposed to be constant regardless of the animals’ age. However,
a deficiency in amino acids not only impacts muscle growth but
also muscle composition. A restriction in amino acid intake (Lys,
Met, Thr and Trp) in post-weaned piglets enhanced carcass fat
compared with pigs fed diets not limited in amino acid content. The
composition of growth (lipid/protein deposition) was altered by
amino acid levels, generating differences in body composition
(Martinez-Ramirez et al., 2008). Similarly, Conde-Aguilera et al.
(2014) evaluated the effects of a 16% deficiency in sulfur amino
acids on body amino-acid composition and carcass quality in
growing-finishing pigs. After 17 weeks, the final body weight was
reduced by 12 kg in deficiently fed animals, as was carcass and
muscle weight. In addition, the animals responded to a deficient
TSAA supply by reducing the protein content of Longissimus dorsi
muscle (a glycolytic muscle) and the Rhomboideus muscle (an
oxidative muscle). The reduction in protein content was moderate
for both muscles (6%), which is smaller than the 20% reduction in
protein content of Longissimus muscle observed by Conde-
Aguilera et al. (2010) during a severe TSAA deficiency in weaned
piglets. As others have observed, the reduction in muscle protein
content is likely accompanied by an increase in lipid deposition
(Castellano et al., 2015; Conde-Aguilera et al., 2010). Similar
results were observed in broilers, in which a 20% deficiency in
TSAA resulting in lower protein gain and an important increase of
lipid gain in the pectoralis muscle (+78%), abdominal fat (+28%)
and, to a lesser extent, in the carcass (+10%). A large fraction of

the amino acids in birds offered a deficient diet could not be used
for protein deposition. These amino acids would then be
deaminated and the carbon chains used for lipid deposition, thus
explaining the higher lipid content. Moreover, the deficient TSAA
supply resulted in an increased Met content in liver protein but a
reduced Cys content in the Rhomboideus muscle, distal jejunum
and ileum. It seems that the intestines and liver may be given
priority during a nutrient deficiency, thus reducing the amino acids
available for protein deposition (Conde-Aguilera et al., 2014).
In some cases, amino-acid deficiency can also affect the
amino-acid profile of the muscle. According to Conde-Aguilera et
al. (2015), a limited TSAA supply affected the concentration of
amino acids such as Met, His and the branch chain amino acids in
muscle protein. A greater His concentration was observed in the
longissimus muscle and semitendinous muscle (a muscle of the
hind leg involved in locomotion) in pigs fed with the deficient Met
group compared to the adequate Met group. The greater His
concentration in deficient TSAA fed−piglets may be indicative of a
greater protein breakdown. Conversely, Chang and Wei (2005)
reported that the amino-acid composition of four different muscles
was not changed in pigs fed a Lys-deficient diet. The lack of
response may have been due to the moderate AA deficiency
applied. Moreover, the redox status of the muscle can be affected
by amino-acid deficiency. Total glutathione (GSH) and reduced
glutathione concentrations were lower in deficient TSAA−fed pigs
than in adequate TSAA−fed pigs, especially in the Rhomboideus
and semitendinous muscles, which are oxidative muscles. The
glycolytic longissimus muscle was not affected (Conde-Aguilera et
al., 2015). Because Met is the precursor of Cys synthesis, the
lower antioxidant status may be the result of a limited Cys supply
for GSH synthesis (Metayer et al., 2008). Cys and GSH act as
chain-breaking elements of the oxidative processes, the deficiency
of which may produce deleterious effects such as lipid and protein
oxidation in the meat.
NUTRITIONAL STRATEGIES TO IMPROVE PROTEIN

DEPOSITION AND MEAT QUALITY IN BROILERS AND PIGS
Nutritionists’ objective is to optimize growth and tissue

accretion by adjusting nutrient density such as amino acids. The
early phase is considered a window opportunity to affect muscle
growth. However, most studies have demonstrated that feed
restriction during this period is damaging to muscle accretion
(Velleman et al., 2014). In piglets, the parenteral administration of
leucine pulses during continuous feeding has been shown to
increase muscle protein synthesis in neonatal pigs (Boutry et al.,
2013). However, only dietary nutritional strategies allowing
increases in muscle growth and improvement in meat quality will
be reported here in. Recent studies have demonstrated that
providing dietary amino acids above the requirement for growth
performance leads to improvement in meat-quality traits. Berri et
al. (2008) have measured the response of broilers to increasing
dietary Lys concentrations from 21 to 42 d of age. An increase in
dietary true digestible Lys from 0.83 to 0.93% resulted in an
increased growth rate, improved feed conversion and increased
breast-meat yield. Body composition traits were not significantly
improved above 0.93% dig. Lys. Final breast pHu increased from
0.83 up to 1.03% dig Lys, and drip loss correlatively decreased
(Figure 3). Strong evidence exists that as pHu increases, the WHC
and potentially the processing ability of broiler breast meat are
improved (Le Bihan-Duval et al., 2001; Zhang and Barbut, 2005).
Figure 3 - Impact of Lys level on meat yield and meat-quality

characteristics in broilers fed from 21 to 42d (Berri et
al., 2008).
Over even a short-term period (3 days) before slaughtering,

Guardia et al. (2014) demonstrated that it is possible to modify the
pHu and color of broiler breast meat. Diets varying in Lys, other
amino acids and energy content were offered to broiler chicks from
36 to 42 d of age. They consisted of a control diet with adequate
amounts of amino acids other than Lys; 6 diets isocaloric to the
control diet, including 3 Lys-deficient diets with an adequate, low,
or high amount of other essential amino acids and 3 Lys-rich diets
with an adequate, low, or high amount of other essential amino
acids. Whereas breast meat yield and abdominal fat percentages
did not differ, the pHu and color parameters of breast meat were
significantly affected by changes in amino acids but not by
changes in dietary energy content. Although the changes in pHu
caused by variations in dietary AA profiles were fairly moderate,
they significantly affected breast meat lightness (Guardia et al.,
2014). In another study, the interaction of Lys and Met levels was
assessed to determine the effect of respective amino acids on
broiler breast-fillet development and meat quality (Zhai et al. 2016

a & b). The experiment compared the effect of increasing dose of
digestible TSAA (from 0.54 to 0.76%) during the 21–42 days
growing-finishing period at two Lys levels (0.89 and 1.09). The
results showed that a higher Lys level allowed a significant
positive effect on breast-fillet development. At a high Lys level, the
TSAA level showed a significant linear effect on breast-meat
development. Moreover, meat-quality aspects were also tested
and demonstrated that a lower cooking loss was obtained with
higher dietary levels of Lys and TSAA (1.09 and 0.76). Zhai et al.
(2012) also observed the effects of Met on the regulation of
cellular processes through proteomic expression in breast muscle.
They revealed that dietary Met levels play a role in different
metabolic pathways such as citrate cycle, actin cytoskeleton
signaling, calcium flux signaling and clathrin-mediated
endocytosis. These modifications concurred to lower the energy
metabolism, modify the ratio of actin to myosin and increase the
nutrient absorption of the muscle, resulting in the specific
development of breast muscle. The higher dietary level of both Lys
and Met resulted in the modification of the ratio of actin to myosin,
which could be linked to the effect observed for cooking losses.
These studies indicated the specific effects of amino acid
on meat quality that belong to signaling function of amino acids
more than the regulation of protein accretion. Gondret et al. (2017)
recently conducted a trial in growing-finishing pigs in which a
short-term supply of high doses of Met (3 times: Met+ and 5 times
the requirement: Met++ treatment) over14 days before
slaughtering improved the pHu and meat antioxidant potential.
Moreover, the dietary supply of high doses of Met as OH-Met (or
HMTBA) showed no detrimental effects on growth performance
and carcass yields. Muscle protein content was not affected, but
lipid content was reduced with an increased dose of Met. In
addition, the highest dose of Met led to an increased level of GSH
in the muscle, thus enhancing its capacity to cope with reactive
oxygen species (ROS). pHu was higher with the highest dose of
Met, which supports the trend of reduction of drip losses and

defrosting losses obtained with this dose of Met. Meat color was
less pronounced and evolved less during storage. Altogether,
these results contribute to improving the meat quality index (Figure
1) and thereby the ability of the muscle to be processed in ham.
Figure 4 - Impact of Met dose on meat quality index in pork

(Gondret et al. 2017)
The meat quality index (MQI) is a predictor of the ratio

between weight of cooked ham to weight of defatted and boneless
fresh ham and was calculated using the equation MQI=−41+11.01
pHu SM − 0.231 L*GS + 0.105 WHC GS (with SM:
semimembranosus muscle, GS: gluteus superficials, WHC: water
holding capacity) (Tribout and Bidanel, 2000).
OXIDATIVE PROCESS DURING MEAT STORAGE
Redox reactions are well known as modulators of metabolic

pathways, as the oxidized or reduced status of a given molecule
can be compared to an electric switch with two positions: “on/off”
(Toledano et al. 2007). It is also well known that cells are equipped
with different antioxidant systems that can directly scavenge free

radicals (e.g., superoxide dismutase (SOD) that detoxify O2°-
radicals) and vitamins that can detoxify oxidized products (e.g.,
vitamin E stops the elongation chain of lipid oxidation). However,
all of these scavenging systems need to be supplied directly
(vitamin E) or require specific nutrients/minerals to be synthetized
by the cell (Cu and Zn for SOD, Se for selenoproteins, sulfur
amino acids for glutathione synthesis). This is currently accepted
to living animals, but when discussing redox processes in meat,
the concept appears to be different because the building blocks
needed to synthesize these components are no longer supplied
and because new synthesis of antioxidant enzymes is impossible
after death. Moreover, meat is prone to producing free radicals
such as ROS (Cadenas & Davies, 2000) or reactive nitrogen
species (Zhang et al. 2013) that can attack the constituents of the
muscle such as pigments, lipids and proteins. The more obvious
example of these oxidative processes is the change of color in
beef meat that appears bright red when the myoglobin is under an
oxygenized status (Fe2+=O) and turns to a brownish/greenish
color when oxidized (Fe3+) (Renerre & Labas 1987). However, the
oxidative processes in the meat are not limited to myoglobin, and
other muscle meat constituents like lipids and proteins, are also
damaged by these oxidative events. Lipid oxidation in meat is
certainly the most studied, as the alteration of lipids by free
radicals can lead to warm over flavor after cooking. The oxidation
of the phospholipids in the membranes will result in membrane
disorganization and cell compartments breaking, allowing passive
diffusion and reaction of the different constituents of the muscle.
Moreover, lipid oxidative intermediates like alkyl or peroxyl radicals
can also spread the oxidative process to neighbor constituents
and then propagate the oxidation process. Muscular proteins are
also targets of free radicals in meat during storage, with
differences depending on the amino acid considered. For instance,
Cys and tyrosine side chain oxidation will lead to intra- or
interprotein bonds through the formation of disulfide or dityrosine
bridges that modify the protein conformation often leading to

functionality losses. Basic amino acid oxidation, such as Lys, Arg
and Pro, will result in carbonyl formation on the side chain, which
can affect protein properties such as WHC and/or gel forming that
would further affect the processing rate and product properties
(e.g., cohesiveness, texture). After slaughter, deboned and piece-
cut muscle are directly exposed to air with a partial pressure of
oxygen of 20%, whereas in a physiological configuration, the
oxygen pressure in the muscle is 5%. Handling, lightening,
grinding, cooking and modified atmosphere packaging are also
pro-oxidative inducers that will enhance free-radical production in
meat, leading to the oxidization of meat components during
storage. Thus, it appears that animal antioxidant potential at
slaughter is the only endogenous reserve allowing fighting more or
less longer with pro-oxidative events. Renerre et al.(1996)
demonstrated that in beef oxidative muscles (the psoas major or
diaphragma), the activity of antioxidant enzymes such as SOD and
catalase was higher than in glycolytic muscles (longisimus
lombarum). Conversely, the color stability is better and oxidative
processes lower in glycolytic muscle compared to oxidative
muscle, suggesting that though the protection appears to be
higher in oxidative muscles, it is not sufficient to ensure stability
during postmortem storage. Moreover, supplying a high quantity of
dietary vitamin E and/or selenium decreases the impact of
oxidative processes during meat storage (Mercier et al. 1998;
Surai, 2002) confirming that nutritional strategies can improve
meat preservation.
Even though only a few examples exist of the effects of
amino acids on oxidative processes during refrigerated storage or
further processing, all examples are linked to sulfur amino acids
and related compounds. TSAA are responsible for glutathione
level and status (GSH/GSSH). Moreover, OH-Met was
demonstrated to be better transsulfurated than DL-Met (Martin-
Venegas et al., 2007),resulting in lower postmortem oxidative
processes under different conditions (Willemsen et al., 2011;
Swennen et al., 2011, Tang et al., 2011). Berri et al. (2012),

comparing the effect of Met sources on broilers’ meat quality traits
during storage, showed that OH-Met allowed lower lipid oxidation
after 12 days of refrigerated storage of breast muscle (pectoralis
major) and thigh muscle (gracilis) during storage (Table 1).
Table 1- Breast and thigh muscle lipid oxidation measured as

TBA-RS according to dietary treatment after 12 days of
refrigerated storage (Berri et al.,2012)
TBA-RS p Value
Muscle type Dietary Met
(mg of MDA/kg of meat) Met source
Breast DL-Met 2.02
Pectoralis major HMTBA 1.16 P=.10
50/50 1.94
Thigh DL-Met 3.85 a
Gracilis HMTBA 2.70 ab P=.02
50/50 2.53 b
MEAT DEFAULTS IN POULTRY: WOODEN BREAST AND

WHITE STRIPPING
As the most valuable meat portion from broiler chickens,

breast meat must meet high-quality market-presentation
standards. Therefore, it is of great importance to breeding
companies and broiler producers that breast muscle growth is
monitored to give the white meat the quality expected from it.
Recent reports of increased cases of breast-muscle myopathies
have brought concern to the broiler-meat industry because
affected carcasses can be downgraded or more frequently
condemned, leading to economic losses (Bailey et al., 2015). This
is the case with the white stripping (WS) and wooden breast (WB)
conditions, which appear to affect only the pectoralis major as
opposed to deep breast myopathy (also known as green muscle or

Oregon disease) (Bailey et al., 2015), which only affects the
pectoralis minor.
White stripping is characterized by white striations
appearing parallel to the direction of muscle fibers in broiler breast
fillets (Kuttappan et al., 2013). Although breast meat that is visibly
affected by WS contains slightly higher levels of fat and may be
visually unappealing to the consumer, no known health or safety
concerns are associated with consuming breast meat with WS.
The etiology of WS is still unknown and is not related to any
specific commercial broiler strains but seems to affect broilers with
higher growth rates and heavier breast weights (Ferreira et al.,
2014).
Wooden breast is characterized by variable degrees of
hardness in the pectoralis major, showing bulging and pale
expansive areas and by variable amounts of interstitial connective
tissue accumulation and fibrosis (Sihvo et al., 2014). Lesions
might start to develop as early as 2 to 3 weeks of age and can
affect a high proportion of birds in a flock, up to 60%, as currently
reported.
High growth rates and enhanced breast-meat yields are
often considered leading causes for the increased presence of WS
and WB in broiler chickens (Petracci and Cavani, 2012; Kuttappan
et al., 2012; Sihvo et al., 2014). However, comparing broiler lines
differing in terms of selection for breast yield indicated a strong
non-genetic component for all the breast muscle myopathy traits
(Bailey et al., 2015). Breast muscles are particularly sensitive to
dietary concentrations of Lys because it represents almost7% of
the total protein content. Dietary Lys thus affects growth and
breast-meat yield. Recent works on dietary Lys have suggested an
influence on the occurrence of WS or WB. However, the main
conclusion of the most recent work Cruz et al. (2016) published
showed that increasing digestible Lys levels resulted in improved
broiler performance to a maximum. Birds with higher bodyweight
also presented higher proportions of myopathies with even
increased severity. As breast weight and growth rate are the direct
resultsof increasing Lyslevels, myopathies do not seemto be
associated with Lysitself but with gains in performance.
CONCLUSION
Pork and poultry meats are often the most affordable meats
around the world. Due to their low lipid content, nutritionists
promote them. Moreover, without any religious constraints, broiler
meat has largely taken the lead in the development of meat
consumption worldwide. With the development of consumers’
desires for top quality meat with nice organoleptic properties and
no hygienic concerns, pressure has developed on poultry and pork
producers to focus increasingly on the quality aspects of those
meats. New trends have appeared, such as meat enrichment in
omega-3 and poly-unsaturated fatty acids through incorporation of
new feedstuffs or extract in the diets. Enriching these meats with
vitamin E and/or selenium with the development of pure organic
forms has shown a beneficial effect on their oxidative stability.
However, dietary amino acids have long been restricted to
muscle growth by itself but not so much to their potential effects on
meat properties and, more largely, meat quality. Recent scientific
works have demonstrated that dietary amino-acid supplementation
or deficiency influence the amino-acid profile of the muscles and
thus the meat properties, influencing oxidative stability, tenderness
and drip loss. Moreover, a short-term dietary supplementation with
amino acids such as Met, OH-Met or lysine appears to be an
efficient solution to improve meat quality without too much
increase infeed cost. More work needs to be done to revise our
amino-acid recommendations according not only to growth but
also to meat quality.
The future shows that broiler meat has tremendous
potential as long as scientists and producers are able to solve the
recent issues of myopathies such as WB and WS linked to the
high growth rate geneticists manage. Years ago, leg abnormalities
were an important issue in broiler production, but science and

genetics took the lead to solve this issue. With a large number of
scientists and approaches focusing on breast myopathies, we can
maintain confidence that a solution will be found.
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BASIC CONCEPTS OF STATISTICS THAT A
NUTRITIONIST MUST KNOW
Fabyano Fonseca e Silva
Universidade Federal de Viçosa, Departamento de Zootecnia
Av. PH Holfs, Campus UFV, Centro, Viçosa, MG, Brazil
ABSTRACT
Usually, experiments related to poultry and swine nutrition

are carried out in situations in which researchers can handle the
experimental conditions. Designed experiments are specified to
consider the partition of the sources of variability into distinct
components in order to examine specific questions of interest.
There are many experimental designs that can be exploited under
this context. One of the basic knowledge of a nutritionist is to
recognize the various experimental designs and use appropriate
statistical methods and software to achieve reliable and relevant
results to be used in practice.
INTRODUCTION
The pig and poultry industries have evolved at a very rapid

rate, and the changes are the consequences of developments in
production technologies driven by a strong global research and
development sector. In other words, these industries have a strong
interest in science and utilize experiments as an important basis
for management decisions (Patience, 2016).
Experimental statistics is a very important feature for
research trials in the field of nutrition of poultry and swine. The
correct planning of experiments and posterior data analysis trough
suitable statistical methods are very important to ensure that the
obtained results are reliable and useful in practice.
Statistical techniques should be considered as research
tools that can produce meaningful, reliable, and unbiased results
when properly applied to situations for which they are designed.
However, no statistical technique can protect against poor

planning, inaccuracies in the data, unsound analysis, or incorrect
interpretation. High-quality research requires proper planning and
careful execution of experiments, correct application of statistical
techniques, and, finally, interpretation of results by one who
understands not only the statistical techniques, but also the field to
which they are applied (Aaron and Hays, 2001).
In summary, the following steps must be taken into account
when designing an experiment: define the problem and the
questions to be addressed (for example, digestibility or growth
trials); define the population of interest (for example, nursery or
finishing); determine the scheme of data collection and sampling
(for example, number of pens and number of pigs per pen); define
the experimental design (for example, completely randomized -
CR, or randomized complete block design - RCB).
After having chosen the suitable protocol to collect the data,
they have to be analysed with appropriate statistical methods
defined according to the characteristics of the evaluated variables
(continuous values, ratio measures, binary or count data) and the
employed treatments (categorical or quantitative levels).
POPULATION OF INTEREST, SAMPLING AND DATA

COLLECTION
A population is a collective whole of animals that

researchers collect data from. Before collecting any data, it is
important that researchers clearly define the population, including
a description of the members. The entire population for which the
researcher wants to draw conclusions will be the focus of the
experiment.
The first consideration on population to be used in an
experiment is the availability of resources in terms of financial
support, personnel, animals, equipment, and time. Then, under the
restrictions imposed by the available resources related to the
population, experiments should be designed.
Other relevant point is the selection of animals from the

whole population to be used in a given experiment. Commonly, it
is referred as sample, which is one of many possible sub-sets of
units that are selected from the population. Under a well-executed
procedure, the results from a sample can be used to draw valid
inferences about the population. A valid sampling must to
considerer information related to how the animals will be assigned
to the experiment and to the dietary treatments, the timing and
methodology of the collection of data and the handling of data and
biological samples. For example, in digestibility trials, special
attention must be given to collect faeces, quantitatively and/or
separately, as required by the assumed protocol. In all instances,
faeces need to be collected free from contamination, especially
with feed (Aaron and Hays, 2001).
A relevant question is to determine how large should be the
sample size of the experiment. Determining the sample size
requires some knowledge of the observed or expected variance
among sample members in addition to how large a difference
among treatments you want to be able to detect. Another way to
describe this aspect of the design stage is to conduct a
prospective power analysis, which is a brief statement about the
capability of an analysis to detect a practical difference (Aaron and
Hays, 2004). This analysis can be realized through SAS (proc
power) and R (package power Analysis) statistical software.
Traditionally, nutrition research has been conducted in such a
manner as to reduce biological variation in order to maximize the
power of the experiment (Festing and Altman, 2002).
According to Aaron and Hays (2004), power calculations
can be made during either the planning or the analysis stage of an
experiment. In either stage, essential information includes
significance level, size of the difference or effect to be detected,
power to detect the effect, variation in response, and number of
replications or sample size. Although there are specifics formulas
and sophisticated algorithm to estimate the power of a given
analysis from a given experiment, Morris (1999) observed that it is
frequently the most neglected part of a pre-experimental protocol.

In reality, investigators seem to choose the number of replicates
arbitrarily on the basis of cost or availability of animals, housing
considerations, convenience, or tradition.
Using specific formulas based on theoretical statistical
criteria, Aaaron and Hays (2004) realized several power analyses
in pigs. The authors commented that in deciding on the minimal
treatment response to be detected, an investigator must consider
what might be biologically or economically meaningful. This is
often difficult to ascertain and depends on the specific situation;
however, the minimal treatment effect must be large enough to
make the experiment worthwhile biologically and/or economically.
As the size of the difference to be detected decreases, the number
of replications required for an experiment of known power will
increase. In this context, these authors proposed a general table
(Table 1) from which it is possible to estimate the number of
replications in a CRB design for litter size based on the
significance level, coefficient of variation and expected differences
in the treatment means (measured as % of the mean).
Table 1 - Number of replications in a CRB design for sow litter

size. Adapted from Aaaron and Hays (2004)
Experimental power is an indication of the probability that

an experiment will arrive at a proper conclusion (Zar, 1981; Baker-
Bausell and Li, 2002; Aaron and Hays, 2004; Shim and Pesti,
2012; Pesti and Shim, 2012). According to Pesti et al. (2016), the
detectable difference is a measure of experimental power, but
there are certainly no guarantees that the appropriate conclusion
will be reached from one experiment. Increasing experimental
costs (number of replicates per treatment and birds per replicate)
merely increases the odds that appropriate conclusions can be
made. These authors presented a graph to illustrate (Figure 1) the
detectable differences between treatment levels in function of the
number of repetitions and the number of animals (in this case,
birds) by pen.
Other term related to power analysis in experimental
statistics is the sensitivity. According to Aaaron and Hays (2001),
it refers to the ability of the experiment to detect real differences, if
they exist, at the desired probability level. The terms power,

precision, and sensitivity have very similar meanings. Given two
experiments, the one that can detect the smaller difference
between two means has the greater sensitivity (power or
precision). Size of the experiment, as measured by the number of
replications, is highly dependent on desired sensitivity. In turn,
sensitivity depends on size of the difference to be detected and
size of experimental error. The chance of detecting smaller
differences
The data collection plan must be done aiming to provide the
maximum amount of information that is relevant to a problem
given the available resources. Understanding how the relevant
variables fit into the design structure indicates whether the
appropriate data will be collected in a way that permits an
objective analysis that leads to valid inferences with respect to the
stated problem. Before collecting the data, it is relevant to take
time to explain the data collection procedures to persons working
in the experiment. Additionally, it is equally important to double
check that all the instruments are valid, reliable, and calibrated.
Figure 1 - Illustration of the detectable differences between

treatment levels in function of the number of repetitions
and the number of animals (in this case, birds) by pen.
Source: Pesti et al. (2016).
Experimental designs
One of the main goals of a designed experiment is to

partition the effects of the sources of variability into distinct
components in order to examine specific questions of interest. The
objective of designed experiments is to improve the precision of
the results in order to examine the research hypotheses. The most
used designs in nutrition research include the CR and the RCB,
being the difference between them characterized by the process of
allotting treatments to experimental units (EU). In general, the

experimental unit is the smallest division of experimental material
to which a treatment can be assigned.
According to Aaron and Hays (2001), the correct definition
of the experimental unit is important because it is the variation
among experimental units treated alike that provides the unbiased
estimate of error for evaluating treatment effects. An experimental
unit may be an individual (e.g., pig) or a group of individuals (e.g.,
litter, pen). Whether an individual or a group of individuals, it is
important that researchers understand the experimental unit is the
smallest entity receiving a single treatment, provided two such
entities could receive different treatments.
The CR is the simplest of all designs since assumes that
the treatments are allotted to EU independently of any factors
(Figure 2). This design allows for the most degrees of freedom for
the residual term in the model to test for treatment significances.
However, it can be unreliable if the EU are not homogenous. Non-
homogeneity of EU can cause inflated error variance components
and can increase the chance of a type 2 error (occurs when the
null hypothesis is false, but erroneously fails to be rejected). In the
RCB, treatments are allotted to EU on the basis of some factor,
commonly referred to as the blocking factor, which should reduce
the error variance if the blocking factor is important. The blocking
factor groups EU based on that particular factor into a block, with
each treatment having a minimum of one EU in each block (Figure
3). The primary function of blocking is to obtain groups of
homogenous EU. Blocking factors vary according to the type of
trial and may be different depending on the desired treatment
structures.
The correct definition of the experimental designs makes
the desired statistical analyses possible, and improves the results
by minimizing the influence of the uncontrolled factors. These
factors are denominated as experimental error, and refers to the
natural, random variation expected under repetition of the
experiment. In other words, it is a collective term often used to
describe variation resulting from all sources of variation

unaccounted for in the experiment.
Experimental error is the difference between a
measurement and the true value or between two measured
values. Experimental error, itself, is measured by its accuracy and
precision. Accuracy measures how close a measured value is to
the true value or accepted value. Since a true or accepted value
for a physical quantity may be unknown, it is sometimes not
possible to determine the accuracy of a measurement. Precision
measures how closely two or more measurements agree with
other. Precision is sometimes referred to as repeatability or
reproducibility A measurement which is highly reproducible tends
to give values which are very close to each other.
Figure 2 - Scheme of completely randomized design considering

five treatments (for example, five different diets) and six
replications (admitting homogeneity of the
experimental material and/or location where the
experiment was installed).
Figure 3 - Scheme of a randomized complete block design

considering five treatments (for example, five different
diets) and six blocks (for example, six differences
locals into a research facility, or six different classes of
body weight at the beginning of the experiment).
Both CR and RCB design consider the two main principles

of the experimental statistics, the replication and the
randomization. Under a general viewpoint presented by Aaron and
Hays (2001), the replication refers to the assignment of more than
one experimental unit to the same treatment. The role of
replication is to provide an estimate of experimental error, so that
increasing the number of replications it is expected a decreasing in
the experimental error. On the other hand, the randomization
refers to the assignment of treatments to experimental units so
that all units have an equal chance of receiving a treatment, thus
ensuring that estimates of treatment means and differences are
unbiased. Additionally, the RCB design presents an extra principle
called local control, usually denominated as blocks. In summary,

they are groups of experimental units that are formed to be as
homogeneous as possible, and improves the comparison of
treatments by randomly allocating levels of the treatments within
each one of these blocks.
STATISTICAL ANALYSIS FROM EXPERIMENTAL DESIGNS
There are several of experimental design, and each one

can be analyzed by using a specific analysis of variance (ANOVA)
that is designed for that experimental design. Nutritionists must
recognize the various experimental designs and to analyze the
data from these experiments by using appropriate methods and
software.
In statistics, an effect is defined as a change in the
response due to a change in a factor level. There are different
types of effects. One objective of an experiment is to determine if
there are significant differences in the responses across levels of a
treatment (a fixed effect) or any interaction between the treatment
levels. If this is always the case, the analysis is usually easily
manageable, given that the anomalies in the data are minimal
(outliers, missing data, homogeneous variances, unbalanced
sample sizes, and so on). Inferences for how the analyzed
variables respond to a given treatment (i.e. inference on effects´
significance) is usually based on hypothesis tests. One additional
consideration in the statistical analysis of experiments is the
situation involving two or more factors, since the challenge is to
differentiate whether the levels of the factors are interacting and
how to slice this interaction in order to produce results with
practical interest.
Generally, hypotheses are stated in a form that is believed
to be untrue (null hypotheses), and data from the experiment are
used to reject or disprove this hypothesis. In this way, the null
hypothesis acts as a base from which an alternate hypothesis may
deviate. According to the example provided by Aaron and Hays
(2001), for a swine nutrition experiment, a null hypothesis can be

given by “the growth rate of pigs fed the normal corn diet is equal
to the growth rate of pigs fed the high-lysine diet.”; whereas the
alternate hypothesis can be given by “the growth rate of pigs fed
the high-lysine corn diet is different than the growth rate of pigs fed
the normal corn diet.” The decision to reject the null hypothesis in
favor of the alternate is based on a statistical test of significance,
in this case an ANOVA F test.
According to Patience (2016), in livestock experiments the
first decision is the selection of ANOVA or regression analysis.
Additionally, a derivative of ANOVA is the analysis of covariance
(ANCOVA), which corrects for potential bias in the data. For
example, initial body weight is often used as a covariate in growth
studies, since it is well known that initial body weight can affect
growth rate, feed intake and feed efficiency, independent of the
experimental treatments. If analysis of covariance is to be used in
this instance, it is critical that the range in initial body weights is
such that there is considerable overlap in all treatments. If such
overlap does not exist, it is not possible to differentiate differences
in performance outcomes between those due to treatment and
those due to differences in the covariate.
There are some assumption to be assumed in the
experimental data analysis by using ANOVA, among the them the
following are highlighted. The error terms are assumed to be
normally and independently distributed with zero mean and a
common (homogeneous) variance. Violation of these assumptions
may affect both probability levels and precision. However, minor
violations are usually not serious (Aaron and Hays, 2001). As a
precaution, however, assumptions should be checked to ensure
that major violations do not exist in the data being analyzed.
Other relevant assumption is related to independence of
data collected from EU. Commonly, if the randomization is
correctly realized in the experiment, the assumption of
independence is verified. However, when the data are collected
under the same EU over time, there is a special situation in
experimental statistics called “repeated measures analysis”.

Because the performance of the animal in time period two will be
at least somewhat dependent on their performance in time period
one, this kind of dependence must be accounted in the analysis
(Patience, 2016). Usually, it is done by assuming special classes
of residual covariance matrices under a repeated measures
approach. It this kind of analysis is not considered when
substantial dependences occurs across time periods, unrealistic
conclusions can be observed. More description about this method
is presented below.
REPEATED MEASURES ANALYSIS
The repeated measures experiment is a common design in

animal science research (Goonewardene et al. 2000; ZoBell et al.
2003). According to Wang and Goonewardene (2004), this
analysis refers to multiple measurements made on the same
experimental unit, observed either over time or space. In repeated
measures designs, the usual practice is to apply treatments to
experimental units and measurements are made sequentially over
time. With this type of experimental design, there are basically two
fixed effects (treatment and time) and two sources of random
variation (between and within animals). Some of the more
common designs in animal sciences include repeated
measurements of such things as weight, gain, blood parameters,
and products of metabolism and digestibility of nutrients. Such
measurements are commonly taken on subjects which have been
randomly allocated to fixed treatment effects such as feeds, drugs,
hormones, etc., with pens or blocks considered as random effects
in the design (Silvia et al. 1995; Wells and Preston 1998;
Goonewardene et al. 2000; Platter et al. 2003).
In general, measurements evaluated on the same animal
are more likely to be correlated than two measurements taken on
different animals, and two measurements taken closer in time on
the same animal are likely to be more correlated than
measurements taken further apart in time. According to Wang and

Goonewardene (2005), he basic objectives for repeated measures
data are to examine simple factor effects (main effects) and the
interaction effects between them. The distinguishing characteristic
of the repeated measurements analysis model from other models
is the assumption about the error variance and covariance
structure (Wolfinger 1996; Littell et al. 1996; 1998; Templeman et
al. 2002). With the repeated model, the usual assumptions about
error variances being independent and homogeneous are no
longer valid (Wolfinger 1996; Littell et al. 2000). The analysis of
repeated measures data therefore requires an appropriate
accounting for correlations between the observations made on the
same subject and possible heterogeneous variances among
observations on the same subject over time. There are different
matrices to model the these (co)variance values over time. A brief
description about these matrices are presented here.
The unstructured matrix is the easiest to understand, but
most complex to estimate. It means that are not imposed
constraints on the (co)variance values. For example, if we had a
good theoretical justification that all variances were equal, we
could impose that constraint and have to only estimate one
variance value. In an unstructured covariance matrix there are no
constraints. Each variance and each covariance is estimated
uniquely from the data, thus resulting in the best possible model
fit, because each variance and covariance values is very close to
what the data reflect. However, it has a cost in terms of the
number of unknown parameters to be estimated implying in many
degrees of freedom and consequently some related convergence
problems. On the other hand, many times there are patterns in
these variances and covariances. For example, all the variances
may be nearly equal, and the covariances may be nearly equal. In
this case, the so called compound symmetry structure is
recommended, This matrix imposes the mentioned constraints,
would “save” a lot of degrees of freedom with little loss of fit,
because it is necessary to estimate only one variance and one

covariance.
There are two ways to run a repeated measures analysis.
The traditional way is to treat it as a multivariate test, where each
response is considered a separate variable. The other way is to it
as a mixed model. While the multivariate approach is easy to run
and quite intuitive, there are a number of advantages to running a
repeated measures analysis as a mixed model.
Missing Data: if the percentage missing is small and the
missing data are a random sample of the data set, this is a
reasonable approach. In the multivariate approach, if a animal is
missing one time point, they will be dropped from the entire
analysis. In the mixed approach, only that time point will be
dropped. The remaining data will be retained.
Post hoc tests: because of the way the Sums of Squares
are calculated in the multivariate approach, post-hoc tests are not
available for repeated measures factors. They are available,
however, using the mixed approach.
Flexibility in treating time as continuous: depending on the
design of the study, rather than consider time as four categories, it
can be more accurate to treat time as a continuous variable. This
allows you to model a regression line for time, rather than estimate
four means.This is not possible in the multivariate approach, but
simple in the mixed approach.
MULTIPLE COMPARISON TESTS
After ANOVA (simple context or under a repeated measure

approach), since the F test has indicated significance for the factor
“treatment”, the researcher may still need to understand subgroup
differences among the different levels of the treatments. According
to McHugh (2011), the subgroup differences are called “pairwise”
differences. ANOVA output does not provide any analysis of
pairwise differences, so how shall the researcher investigate
differences among the various subgroups tested with ANOVA?
The first approach that comes to mind is to perform a number of t-

tests between each of the pairs of interest. This is not a good
approach for two reasons: First, doing repeated statistical tests on
the same data – which is what performing t-tests on each pair of
interest does – causes alpha inflation (1). Second, the results will
still be uninterpretable because individual t-tests can examine only
two groups at a time. Each of the subgroups within an ANOVA has
its own mean. The total number of means (i.e. the count of means,
one for each of the experimental and control groups) is excluded
from analysis when repeated t-tests are used to examine the
pairwise differences from an ANOVA. Ignoring the fact that there
are many more subgroup means in the ANOVA will artificially raise
the number of pairwise differences that are significant, and worse,
the individual pairwise t-statistics will be larger when some
subgroups are excluded from the post hoc testing. Therefore,
using t-tests to examine pairwise differences is likely to
overestimate the size of the individual t-tests. This means that the
sum of t-values from all the pairwise t-tests will often exceed the
value of the t-statistic produced by one of the multiple comparison
analysis statistics. As a result, performing multiple t-tests will lead
the researcher to a higher probability of making a Type I error.
That is, the researcher is much more likely to report significant
differences between some of the pairs that have no real
difference.
Performing multiple pairwise t-tests leads to another
problem. The researcher may wish to test differences between
one or more study groups and a set of combined study groups.
Pairwise t-tests cannot perform that kind of analysis. However,
there are a set of other tests that overcome all the limitations of
the pairwise t-test approach, among them we highlight: Sheffèe,
Tukey, Newman-Keuls and Dunnett.
According to McHugh (2011), each of the multiple
comparison analysis (MCA) tests has its own particular strengths
and limitations. Some will automatically test all of the pairwise
comparisons, others allow the researcher to limit the tests to only
pairs or subgroups of interest. Each approach has implications for

alpha inflation and for the kind of answers the researcher can
derive from the test. Therefore, the choice of an MCA statistic, as
all choices about which statistic to use, should be based on the
specific research questions.
Tukey’s multiple comparison analysis is used to test each
experimental group against each control group. This method is
preferred if there are unequal group sizes among the experimental
and control groups (McHugh, 2011). The Newman Keuls statistic
is appropriate for studies in which even very small differences are
important to find and where the consequences of a Type II (the
failure to reject a false null hypothesis) error are worse than the
consequences of a Type I (the incorrect rejection of a true null
hypothesis) error. This makes it a useful tool for new areas of
science where not much is known about the phenomena of
interest (McHugh, 2011).
The Tukey and Newman-Keuls tests are designed to test
simple comparisons. When the researcher must test subgroups
composed of combinations of experimental and control groups,
other statistics which can test complex comparisons should be
used. The most commonly used statistics in this category are the
Scheffee and Dunnett tests. According to McHugh (2011), The
Scheffee method, tests all possible contrasts, simple and complex.
If it is known in advance that all contrasts are going to be tested,
the Scheffee method is slightly more powerful than all other two
methods. Thus the Scheffee, like the Tukey test, is the more
appropriate test to use when predicted differences are small, and
the consequences of a Type II error outweigh the consequences
of a Type I error. The Scheffee test assumes equal sized
experimental and control groups in the ANOVA. The Dunnett
method is useful for testing control group designs. It is a
particularly powerful statistic and therefore it can discover
relatively small but significant differences among groups or
combinations of groups. The Dunnett method is quite useful when
the researcher wishes to test two or more experimental groups
against a single control group. It tests each experimental group’s

mean against the control group mean. The other methods test
each study group against the total group mean (i.e., the grand
mean). This difference in testing approach makes the Dunnett
method much more likely to find a significant difference because
the grand mean includes all group means and thus mathematically
it is less extreme than individual group means (McHugh, 2011). A
summary of the characteristics of the mentioned test is presented
at Table 2.
Table 2 - General description of the most used multiple

comparisons tests in analysis of experiment. Adapted
from McHugh (2011).
It is therefore important to select the test that best matches

the data, the kinds of information about group comparisons, and
the necessary power of the analysis. It is also important to select a
test that fits the research situation in terms of theory generation
versus theory testing. The consequences of poor test selection are
typically related to Type 1 errors, but may also involve failure to
discover important differences among groups. Multiple comparison
analysis tests are extremely important because while the ANOVA
provides much information, it does not provide detailed information
about differences between specific study groups, nor can it
provide information on complex comparisons (McHugh, 2011).
REGRESSION ANALYSIS
Regression analysis is a form of predictive modelling

technique which investigates the relationship between a
dependent (response variable) and independent variable
(predictor variable affecting the response variable). This technique
is used for modelling and finding the effect relationship between
the variables. There are multiple benefits of using regression
analysis, such as: It indicates the significant relationships between
dependent variable and independent variable;and it indicates the
strength of impact of multiple independent variables on a
dependent variable. There are various kinds of regression
techniques available to make inferences, being the difference
between them driven mainly by the number of independent
variables, the type of dependent variables and shape of regression
line.
Simple linear regression is one of the most widely known
modeling technique. Linear regression is usually among the first
few topics which people pick while learning inference under a
modelling viwepoint. In this technique, the dependent variable is
continuous, independent variable(s) can be continuous or discrete,
and nature of regression line is linear. It establishes a relationship
between dependent variable (Y) and one or more independent
variables (X) using a best fit straight line (also known as

regression line). It is represented by an equation Y=a+bX + e,
where a is intercept, b is slope of the line and e is error term. This
equation can be used to predict the value of target variable based
on given predictor variable(s). Least Square Method (LSM) it is the
most common method used for fitting a simple linear regression
model. It calculates the best-fit line for the observed data by
minimizing the sum of the squares of the vertical deviations from
each data point to the line. Because the deviations are first
squared, when added, there is no cancelling out between positive
and negative values. A special kind of general linear regression is
the polynomial regression. While there might be a temptation to fit
a higher degree polynomial to get lower error, this can result in
over-fitting and lack of biological interpretation, mainly in nutrition
experiments in the field of Animal Science. Always plot the
relationships to see the fit and focus on making sure that the curve
fits the nature of the problem.
While a linear equation has one basic form (straight line or
polynomial curves), nonlinear equations can take many different
forms. It covers many different forms, which explains why
nonlinear regression provides the most flexible curve-fitting
functionality. According to France and Kebreab (2008), animal
nutritionists are primarily involved in the study of the digestive and
metabolic processes regarding the utilization of nutrients. With this
aim, the analysis and interpretation of some response functions
and time-dependent data are of interest to establish the effects of
specific nutrients on processes such as digestion, growth or
lactation, or to examine the time course of events. Usually, this
sort of data follows a curvilinear pattern that may be represented
by a non-linear function.
Non-linear models represent curved relationships and have
at least one parameter appearing non-linearly, i.e. the functional
form of the equation is not linear with respect to the unknown
parameters (Ratkowski, 1983; López, 2008; France and Kebreab,
2008). Non-linear regression is a procedure for fitting any selected
equation to data, endeavouring to smooth data, interpolate for

unknown values and make observations continuous over the
range of values of the explanatory variable (Bates and Watts,
1988; Seber and Wild, 2003, López, 2008; France and Kebreab,
2008). Differently from liner regression, to compute the parameter
estimates in non-linear regression initial values for the parameters
are required. In successive iterations, these initial values are
adjusted so that the residual sum of squares is reduced
significantly in each step. Thus, under this iterative process, new
values for the parameter estimates are continually generated until
a convergence criterion is met (i.e., there are no reduction in the
sum squares in subsequent iterations).
In animal nutrition, the requirement for a specific nutrient
can be defined as the minimal amount of this nutrient (dose)
needed to reach maximum performance (response) assuming that
all the other nutrients are provided in adequate amounts. Usually,
segmented-regression models are fitted to dose-response data in
order to determine the optimal level of a given nutrient.
Segmented regression (or piecewise regression) allows multiple
linear models to be fit to the data for different ranges of the
independent variable (X). Breakpoints, denoted as Xo, are the
values of X where the linear function changes, and when these
points are considered as unknown (as observed in nutrient
requirement analysis), the segmented regression models are
considered as nonlinear.
To infer on true Xo in the segmented regression, the
trajectory of the response (Y) variable (for example, average daily
gain ) over nutrient levels (X) values must be modeling by using
different models, such as quadratic-plateau (QP) model (Fuller
and Gallant, 1974):

a+bXi +cXi +ei , if Xi  X0
2
Yi   ,

 p+ei , if X i  X 0
where: Yi is the response variable measure (for example, average

daily gain); a, b and c are parameters of the quadratic function, X 0
is the true breakpoint; p is the plateau (stabilized value of Y after
K0) and ei is the residual term. As mentioned earlier, the QP
model is nonlinear when X0 is treated as unknown, thus the NLIN
procedure of SAS® software can be used to fit this model using an
iterative least squares procedure based on Gauss-Newton
algorithm. Since X0 is defined into the nonlinear estimation as a
direct function of unknown b and c parameters (K 0=-0.5b/c), its
approximated confidence interval can be obtained by simple "plug-
in" method (Casella and Berger, 1990). Under this approach, it is
possible to estimate the true X0 value as well as to obtain its
confidence interval (as illustrated at Figure 4).
Response variable (for example, average daily gain)
plateau
breakpoint
Nutrient level X0
Confidence interval for X0
Figure 4 - Illustration of a segmented regression fitting through

nonlinear regression approach.
The purpose of this paper is to summarize the basic

concepts of traditional experimental statistics in livestock, focusing
in poultry and swine nutrition. There are many experimental
designs that can be exploited under this context. Each
experimental dataset can be analysed by using a specific ANOVA
that is chosen according to design properties. One of the basic
knowledge of a nutritionist is to recognize the various experimental
designs and use appropriate statistical methods and software to
achieve reliable and relevant results to be used in practice.
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STRATEGIES TO INCREASE PIGLETS
PERFORMANCE AFTER WEANING
José Francisco Pérez, Laia Blavi and David Solá-Oriol
Animal Nutrition and Welfare Service (SNiBA; http://sct.uab.cat/sniba/en ).
Department of Animal and Food Sciences, Universitat Autònoma de Barcelona, 08193, Bellaterra (Spain)
ABSTRACT
Performance of piglets in the nurseries may be variable

depending on the BW of piglets at weaning, but also on the
management, and pathogenic load in the pig facilities. The early
events in the pig life are very important and long lasting as the
bottom pigs will largely impose a significant cost to the system
because of reduced market weights, and increase barns
occupancy. The present report evidences that there is not a
unique solution to improve performance of pigs. On the other
hand, a complex set of dietary and management strategies are
necessary to allow an optimum and efficient growth performance.
Dietary strategies for feeding sows should aim: i.- to enhance fetal
growth (arginine, folate, betaine, vitamin B12, carnitine, chromium,
zinc); ii.- to promote prenatal learning (flavors); iii.- to allow an
early microbial colonization (probiotics); or iv.-to increase milk
production (DL-Methionine, DL-2-hydroxy-4-methylthiobutanoic
acid, arginine, L-carnitine, tryptophan, valine, vitamin E). Dietary
strategies for piglets, either during lactation or after weaning, may
include: i.- to care for an early neonatal colostrum intake; ii.- to
promote early social interactions between litters; iii.- to provide an
early exposure to the new feed (creep feed); iv.- to supply
conditionally essential nutrients (nucleotides, AAs); v.- to reduce
protein fermentation and increase gut digestion (low protein, low
Ca diets, high quality protein ingredients); vi.- to increase feed
intake after weaning (palatable ingredients); or vii.- to look for
zootechnic effects with additives (acidifiers, phytogenic).
INTRODUCTION
Commercial weaning is a sudden change in pig life during

which pigs have to be adapted to eat a novel food after the
separation from their mother. This occurs at d 21 to 28 of age, and
pigs are moved to the nurseries where they pass from a liquid diet
of 20% DM to a compound diet with approximately 90% DM
(Varley and Wiseman, 2001). The nutritional, psychological and
environmental changes produce stress response and anorexia on
the first days after weaning (Pluske et al., 2007). Anorexia
produces gastrointestinal disturbances: alterations in small
intestine architecture and enzyme activities, transiently-increased
mucosal permeability, disturbed absorptive-secretory electrolyte
balance and altered local inflammatory cytokine patterns (Lallès et
al., 2007). The early events occurring at the very young age of
piglets provoke a penalty on the production of nursery, growing
and finishing pigs. The objective of this paper is to present
evidences of negative and positive factors likely affecting the
performance and productive expectation for the growing pigs. The
report evidences that there is not a unique solution to improve
performance of pigs. On the other hand, a complex set of dietary
and management strategies are necessary to allow an optimum
and efficient growth performance.
PERFORMANCE RESULTS IN THE NURSERIES
Performance of piglets in the nurseries may be variable.

Data collected from 700,000 sows in Spain (SIP inform, 2015)
show that, in barns up to 20-22 kg BW in five weeks, average
mortality is approximately 3.3% (from the lowest mortalities found
to be 1.3% to an average highest values of 5.2%) and an average
feed: gain ratio of 1.62 (from the lowest ratios found at 1.45 to an
average highest of 1.78%). The results evidence also variability
among barns during the growing and finishing phase, with a range
of mortalities from 2.4 to 5.2% (averaging 3.8%).
There are a big number of factors affecting performance in

the nurseries. Weaning weight is important to influence post-
weaning growth, morbidity, and mortality. Cabrera et al., 2010
showed that pigs that weigh less than 5.0 kg at weaning represent
the greatest marginal opportunity to decrease days to slaughter (8
d more to achieve 125 kg of BW or weigh 3.3 kg less than 5.5-kg
pigs). The bottom 9% of the pigs (4.1 to 5.0 kg) imposes
significant cost to the system because of reduced market weights,
and increase barns occupancy. The increased time to achieve
target BW is a poor financial option.
Stein (2015) reports also a benchmarking of performance
results that show averages for nursery, finishing and wean-to-
finish closeouts in Canada. For 2013, the dataset included over
17,000 anonymous and confidential closeouts. For average start
weights in nursery groups, there were a huge performance and
mortality penalty for groups with very light (< 4.5kg) average
weaning weights and a substantial a penalty even for groups with
light (5kg) average weaning weights. They also observed that
there is a positive effect of weaning weight on Average Daily Gain,
i.e. the higher the initial BW the higher the Average Daily Gain.
Therefore, the productive results evidence that individual
young piglets bring themselves a source for variation. Variability
may respond to changes that could be related to the intrauterine
growth, birth weight, colostrum intake, microbial early colonization,
or milk yield during lactation. Intra-uterine growth retardation is a
significant problem in pigs. Average litter size of sows has been
increased by genetic selection over the last decade; which it has
been associated with a reduction in the mean piglet birth weight
and, concomitantly, with an increased within-litter variation in birth
weight. As a consequence, this has led to a rise in the proportion
of small piglets (less than one kg birth weight) in large litters
(Quiniou et al., 2002). Pigs with low birth weight exhibited
consistently long-term effects, such as a higher morbidity-mortality
and lower postnatal growth rates, which evidence epigenetic
alterations from the negative effects of dietary restriction during
pregnancy. The growth failure is partly explained by the reduced

intestinal size, and shorter villus height and villus: crypt ratio (Che
et al., 2016), and the lower intestinal trophic responses to enteral
feed introduction during the early postnatal period. Characteristics
of the gut microbiota and binding property of intestinal mucosa to
luminal bacteria, and signs of an immature innate immunity are
also evident in pigs with low birth weight.
However, morbidity and mortality in the barns are also
affected by the sanitary status of the farms. Nowadays health is
defined as the balance between the infection pressure on the pig
and its ability to fight them with its own immune system. Therefore,
the optimization of health in pigs requires a multifactorial
approach.
In one respect, it is necessary to reduce the pathogenic
load in the pig facilities by using proper cleaning and disinfection
protocols, and biosecurity measures. General enhancement of
animal health and welfare to reduce the need for antibiotic use
include, for example, the right management of nurseries with
single origin batches of piglets, high quality water supply, low
density in the pens, or wide space and fresh water in the feeders
to enhance feed intake. In another respect, specific alternatives to
antibiotics are necessary, including vaccination, feeding
approaches and breeding. The present report will focus on some
dietary and feeding strategies which they may help to improve
performance of piglets during the first days of life and after
weaning. Strategies may include a proper feeding of sows during
pregnancy (to promote epigenetic effects in piglets) and lactation
(to increase milk production), and a proper feeding of piglets with
palatable and digestible diets after weaning.
DIETARY STRATEGIES FOR SOWS
Enhancing fetal growth
Pre-weaning piglet mortality is largely attributed to the

incidence of low birth weight and birth weight variation within the
litter. Therefore, developing strategies to increase within-litter
uniformity of piglet birth weight are important (Quesnel et al.,
2014). Dietary supplementation of certain nutrients could enhance
litter size and fetal growth in swine, such as chromium, L-carnitine,
omega fatty acid, lysine, and L-arginine. Piglets born to sows
supplemented with nutrients to increase the birth BW and vitality
are moreover able to suckle for longer, therefore receive more milk
from the sow and grow faster during the suckling period
(Birkenfeld et al., 2006).
Among these studies, strategic Arginine supplementation
to improve placental weight per sow and pregnancy outcomes has
been recognized (Mateo et al., 2007). Arginine is an unusually
abundant amino acid in the conceptus (Wu, 2010a), which it is
utilized by multiple pathways, including the synthesis of protein,
nitric oxide (NO), polyamines, and creatine. Several studies have
demonstrated that polyamines and NO are key regulators of
angiogenesis, embryogenesis, and placental and fetal growth,
especially during the first half of pregnancy (Wu, 2010a). These
changes promote optimal intrauterine conditions to minimize the
losses and improve growth of viable fetuses.
Maternal diet enriched with methylating micronutrients, such
as Folate, Betaine, or Vitamin B12 have been also associated
with increased fetal weight in late gestation (Oster et al., 2016). It
has been reported, these compounds provides beneficial effects
for embryo and placental development, and can increase litter size
in multiparous sows (Lindemann, 1993). Folate, betaine, or vitamin
B12, are integrally involved in methionine metabolism, with
compounds donating monocarbon units for the remethylation of
methionine from the toxic homocysteine.
Supplementing dams with L-Carnitine increased circulating

insulin-growth factor I (IGF-I) concentrations at mid-gestation in
swine (Musser et al., 1999). The IGF are polypeptides that
stimulate proliferation and differentiation of skeletal muscle cells
and are regulators of muscle growth and development. The
supplementation of gestating sows with L-carnitine results in
greater placenta, heavier BW at birth. Additionally, offspring from
sows fed L-carnitine had a larger cross-sectional area and more
total muscle fibers in the semitendinosus muscle than piglets from
controls. An increased maternal IGF-1 concentration, an improved
transfer of glucose from maternal to fetal blood through the
placenta and improved oxidation of glucose, leading to a more
efficient energy production, in the fetuses may be key events
responsible for an improved fetal development.
Lindemann et al. (1995) first reported that diets
supplemented with Chromium from chromium picolinate (CrPic)
increased the number of live pigs born in sows through two
parities. The primary metabolic role of Cr is to potentiate the action
of insulin by facilitating insulin binding to receptors at the cell wall.
Dietary supplementation with CrPic in the diet of sow throughout
gestation increases the body mass gain from insemination to day
110 of gestation and the number of piglets born alive (Wang et al.,
2013).
Fluctuations of Zn status biomarkers may identify critical
periods throughout the reproductive cycle during which high Zn
requirements are not met by dietary Zn intake. It has been shown
that Zn deficiency during gestation may affect the growth and
development of the fetus and induce complications at birth. During
the lactation, mobilization of Zn body stores into milk of sows is
also tightly regulated (McCormick et al., 2014). Plasma Zn is
transferred to the mammary gland, and it provides active transport
for secretion into milk to meet the high Zn requirements of piglets
(Matte et al., 2014). However, we have observed that during the
latter 2 weeks of lactation, sows’ milk is inadequate to meet their
Zn requirements, especially in light pigs that have more risk to
develop Zn deficiency (serum Zn levels below 0.8 at the end of

lactation (Blavi, 2017)
Prenatal Learning
Young animals can learn about volatile compounds

contained in flavors from the maternal diet to stablish feed
preferences which are long-lasting. A learning process by flavors
added in maternal diets before or after birth, known as maternal
learning could smooth the weaning changes and optimize the
production efficiency of pigs. In a maternal-learning process,
volatile flavors coming from maternal diets may reach the fetus via
the amniotic fluid and/or the placental bloodstream (Blavi et al.,
2016). During the last trimester of gestation, fetuses of several
mammalian species appear to be able to detect and retain
chemosensory information. In pigs, it has been observed that pre-
natal exposure to flavors via maternal diet influences feed
preferences of pigs in two-choice test (Figueroa et al., 2013; Blavi
et al., 2016) and enhances their feed intake and growth
performance (Oostindjer et al., 2011; Blavi et al., 2016). The basis
of this effect could be related to the transference of some volatile
compounds to the womb and/or the milk, producing a conditioning
process that may occur inside the womb, or related to an effect of
familiarity due to repeated exposure (Hepper, 1988). Greater feed
intake after weaning could also be related to a stress-reducing
effect by piglet exposure to a familiar flavor acquired before
weaning (Oostindjer et al., 2011). Our results indicate that
conditioning are stronger during late gestation rather than during
lactation, and when the flavor is not simultaneously included in the
creep-feed (Blavi et al., 2016).
Figure 1 - Feed intake of post-weaned piglets in a double choice

between the flavor provided to sows (Fluidarom 1003 ®)
and a control flavor (Lacto-Vanilla). FFF had flavor in
gestation-lactation, creep-feed and weaning; FCF in
gestation-lactation and weaning; and CCC no flavor
inclusion. The number at the top of the bars indicates
the average percent preference for Fluidarom 1003 ® (n
= 6). NS P > 0.1; *P < 0.05.
Probiotics and early colonization
Establishment of the intestinal microbiota after birth plays

an important role in the development of the neonatal gut and its
immune system. It appears that the mother's microbiota is shared
to a large degree by the offspring during the first days of postnatal
life.
Therefore, maternal environmental factors (diet
composition, antibiotic treatment, etc.) that induce changes in
maternal microbiota may have huge effects on offspring gut
physiology (Kelly and Conway, 2005). Recent studies with piglets

exchanged for adoption between two swine breeds demonstrate
the influence of the mothers on the configuration of the microbiota.
The first microbial colonization occurs during childbirth (mother-
belly effect) and evolves in waves of subsequent colonization
during lactation depending on the composition of the milk of the
sow (adoptive mother effect) and the composition of the diet
offered after weaning.
Until recently the environment of the mammalian uterus was
thought to be sterile and so was breast milk. However, recent
studies have shown that the animal's exposure to microorganisms
begins even before birth (Rautava et al., 2012) and that, in
addition, breast milk in humans is capable of delivering certain
intestinal microorganisms from the mother to the offspring using
the mammary via (Rodriguez, 2014). Thus, the mother has some
control over the microorganisms to which the breeding is exposed
in order to determine the correct epigenome programming of the
newborn. In pigs, a positive effect on the microbiota and immune
system of piglets has been obtained from Probiotics included in
the sows feed during gestation (Scharek-Tedin et al., 2013; Baker
et al., 2013).
Increasing the milk production
It is generally accepted that one of the most efficient ways

to stimulate a piglet’s good life start is via the sow milk. Growth of
nursing neonates depends on adequate milk yield from their
lactating mothers. Mammary gland growth, which occurs during
gestation and lactation, is critical to lacto genesis. However, the
main factors conditioning the sow milk production are the number
and vitality of suckling piglets, and the daily feed intake reached by
sows during the lactation curve.
Kim et al. (2009) proposed that increasing mammary gland
growth (including vascular growth) and blood flow to mammary
tissue is an effective strategy to improve lactation performance in
mammals (including sows). Increased consumption of methionine

as DL-Methionine (DLM) or its Hydroxy analogue DL-2-
hydroxy-4-methylthiobutanoic acid (HMTBA) could benefit milk
synthesis and neonatal growth. At postnatal day 14, piglets in the
HMTBA group had higher body weight than those in the Control
group, and tended to be higher than those in the DLM group
(Zhang et al., 2015). Neonatal growth and milk synthesis were
regulated by dietary methionine levels and sources, which resulted
in marked alterations in amino acid, lipid and glycogen
metabolism. Sows fed the HMTBA treatment showed also higher
Lys, Leu, Ile and Val levels, and lower Glu and Gln in the plasma
than those of the DLM-fed sows. Greater plasma levels in sows
fed the HMTBA may be a result of their enhanced net portal
balance (HMTBA doesn´t share the same intestinal transport for
AAs, such as Lys and Met do). The higher plasma AAs level may
explain the higher milk protein synthesis in HMTBA-fed sows than
in Control or DLM-fed sows (Zhang et al., 2015).
As reported during gestation, Arginine is the common
substrate for the generation of nitric oxide (NO; a major vasodilator
and angiogenic factor) and polyamines (key regulators of protein
synthesis). Thus, modulation of the arginine-NO pathway may
provide a new strategy to enhance the growth (including vascular
growth) of mammary tissue and its uptake of nutrients; therefore
improving lactation performance in mammals. In support of this
proposition, supplementing 0.83% L-arginine (as 1% L-arginine-
HCl) to diets of lactating primiparous sows increased milk
production and the growth of suckling piglets (Mateo et al., 2008).
Dietary content of Tryptophan has been related to
variations of feed intake in lactating sows, the mechanisms remain
to be elucidated. Mosnier et al. (2009) have shown that during the
week following parturition, plasma tryptophan and niacin
decreased while plasma kynurenine increased. On the 2nd and
3rd weeks of lactation, plasma tryptophan and kynurenine
returned to pre-farrowing concentrations, while niacin increased
throughout lactation (p < 0.05). This study suggests
that tryptophan catabolism presumably through the kynurenine

pathway is high during the 1st week after farrowing, and that
dietary supply of Niacin and vitamin B6 could be transiently
suboptimal in early lactation.
Branched Chain AAs (BCAA) are spared by the liver and
readily used in tissues with high metabolism. Catabolism of BCAA,
such as valine, occurs in liver, kidney, muscle, heart, adipose
tissue and mammary gland. Recent research with lactating sows
has indicated that Valine is catabolized at a high rate in the
mammary tissue (Li et al., 2009) and it is contained in milk AAs
composition at different ratios (Table 1, adapted from different
sources (King et al., 1993; Wu et al., 1994; Trottier et al., 1997).
Several research has been conducted the last decades to
estimate the correct dietary valine-to-lysine ratio (Val: Lys) for
lactating sows in order to maximize litter performance and
minimize tissue mobilization during lactation. Present standard
ileal digestible (SID) Val: Lys recommendations vary between 70
to 85 % for lactating sows but, experimental trials with positive
effects on piglets weaned per litter have been observed (Tokach et
al., 1993; Richert et al., 1996, 1997a,b; Moser et al., 2000) by the
use of 120%. In contrast, other studies failed to confirm this high
value (Boyd et al., 1999; Carter et al., 2000; Gaines et al., 2006).
A meta-analysis of the available data on lactating sows based on
the effect of Val:Lys ratio on the litter weight gain, confirmed a
0.90:1 Val: Lys ratio recommendation (from Rousselow and Speer,
1980; Richert et al., 1996, 1997a,b Carter et al., 2000 and Gaines
et al., 2006).
Table 1 - Pattern of the different AAs ratios to lysine for different

pools in lactating sows.
Mik Mammary Retained in Plasma

Tissue Mammary gland
Lysine 100 100 100 100
Threonine 59 58 150 86
Tryptophan 77 78 25 69
Methionine 114 116 53 32
Valine 60 58 216 166
Leucine 56 58 403 164
Isoleucine 65 59 231 72
The current NRC (2012) recommendation for SID Val: Lys

is 0.85:1, whereas the Danish recommendation of SID Val: Lys is
0.76:1 (total Val: Lys of 0.80:1; Tybirk et al., 2013). So maybe
thinking in modern hyperprolific sows valine to lysine ratio should
be taken into account when high performance is achieved.
L-Carnitine is required for the transport of medium- and
long-chain fatty acids into the mitochondria for β- oxidation.
Ramanau et al. (2005) described that sows whose diet is
supplemented with L-carnitine produce more milk during lactation
than control sows, even in a strongly negative energy and protein
balance. The study indicates that L-carnitine might also enhance
the utilization of body fat by sows in a strongly negative energy
balance, particularly in primiparous sows.
The addition of vitamin E at high concentration in the sow’s
diet during the last week of gestation and lactation improved the
weight of piglets at weaning, and enhanced humoral immune
function and antioxidant activity in sows and piglets (Wang et al.,
2016). Results showed that supplementation of the maternal diet
with 250 IU/kg vitamin E improved the weaning weight of piglets
and the concentrations of immunoglobulin G (IgG) and
immunoglobulin A (IgA) in sow plasma, colostrum and milk. The
concentrations of fat and α-tocopherol in the colostrum and milk,
and the level of plasma IgG, IgA, total antioxidant capacity were all
also higher in piglets from sow fed vitamin E (P < 0.05).
DIETARY STRATEGIES FOR PIGLETS: THE LACTATION PERIOD
Caring piglets during the first days
For piglets, birth marks a sudden decrease in body Tª as

the starter signal to boost their own thermogenesis. However,
piglets are born with very small energy depots. Without brown fat,
the glycogen depots supply energy in amounts adequate for ∼16 h
(Theil et al., 2011), and energy supplied from colostrum must
contribute to ensure piglet survival because transient milk is not
being secreted until ∼34 h postpartum. In these conditions,
newborn piglets are in a negative energy balance immediately
after birth because of their high physical activity and high energy
need for thermoregulation (Theil et al., 2014). The young piglets
rely on colostrum, and if the amount of colostrum is small, the
piglet may die because of hunger or because it eventually is
crushed by the sow when the piglet is too weak. Colostrum is also
a source of very digestible nutrients and various forms of bioactive
compounds such as immunoglobulins, hydrolytic enzymes,
hormones, and growth factors to boost the small intestine
architecture during the first hours and days (Zabielski et al., 2008).
An intake of 200 g of colostrum per piglet, or 180 g/kg birth weight,
seems to be a minimum requirement to reduce the risk of mortality
(Le Dividich et al., 2005). An average intake of 250 g of colostrum
is recommended to achieve a good BW gain (quantities predicted
as per Devillers et al., 2004; Theil et al., 2014).
Newborn piglets from the same litter (whose number of live
born piglets may easily exceed number of functional teats)
compete for mammary glands, preferably the anterior and the
middle ones. The posterior mammary glands can produce less
beneficial proteins than anterior ones (Wu et al., 2010b). Such
circumstance makes piglet colostrum supplementation a wide
extended practice in pig production, especially to support the low

BW piglets.
Social Interventions
In modern pig production systems, social interactions

between piglets (apart from cross-fostering) are uncommon or
negligible during the suckling period. However, at weaning they
are grouped by BW categories and sexes. Figueroa et al. (2012)
determined the effects of social interactions between litters before
weaning on maternal recognition and how these interactions may
affect performance and behavior of piglets at the suckling period
and after weaning by comparing early socialized litters (48h after
birth) with individual litters for the entire lactation period (28d). It
was observed that during lactation, most early socialized piglets
(94% ± 10%) suckled from their own mother. During the post-
weaning period, the non-socialized piglets showed more
aggressive interactions and a lower occurrence of positive social
interactions (Figure 2) and the prevalence of severely wounded
piglets were higher. Early socialized piglets also improved weight
gain during lactation and the first week post-weaning (7.5% and
20%, respectively).
Figure 2 - Early socialized (CG) and No-socialized (NCG) group

during lactation and their prevalence of interactions
(Negative or positive) and injuries (sever and moderate)
after weaning. Asterisks indicate prevalence differences
between groups (* P < 0.05; ** P < 0.005).
Similar protocols recently conducted in the same research

group by Jordà et al. (2015) with larger number of sows, litters and
piglets showed higher piglet BW for piglets early socialized than
for those raised in commercial conditions (individual litters and
grouped after weaning by sex and BW category) at weaning, 14
days after weaning, and at the end of the nursery period (35d
post-weaning). These studies show that allowing contact between
litters before weaning may facilitate early interaction between
conspecific animals increasing their welfare due to a reduction of
social stress during the post-weaning.
Creep Feeding
Creep-feed are highly palatable and easily digestible diets

that are offered to lactating piglets after the first week or ten days
of lactation. They are always formulated as complex diets but may
vary by using high palatable ingredients (Solà-Oriol et al., 2009a,
b, 2011) combined with different technological processing. Creep-
feeding is one of the most common earliest feeding strategies in
solid feed to promote a suitable transition at weaning and may

contribute to a reduction of pig body weight variability from
weaning and onwards (Solà-Oriol and Gasa, 2016). Creep-feed it
is especially beneficial for the piglets raised in large litters and long
lactations as with the present hyper-prolific sows occurs.
Moreover, it has been reported that eater piglets of the creep-feed
may become early familiarized with the solid diet and they start
early to consume after weaning (Figure 3; Bruininx et al., 2002).
Figure 3 - Percentage of weanling pigs (eaters, the thin line; non-

eaters, the thick line; and no-feed pigs, the dash line)
that had not eaten after weaning (Bruininx et al., 2002).
It is important to remark that not all piglets consume creep-

feed within a litter; approximately only 60% of pigs are creep-feed
eaters (Sulabo et al., 2010; Barnett et al., 1989). Consumption of

creep-feed during lactation stimulated food intake and growth after
weaning (higher ADFI and ADG) during first days of post-weaning
(Bruininx et al., 2002, 2004; Cartensen et al., 2005; Sulabo et al.,
2010) and increases total BW gain (Kuller et al., 2007; Sulabo et
al., 2010) in eaters piglets.
Nutrients: Nucleotides, Amino- acids
The nutritional support of gastrointestinal growth and

function is an important consideration in the clinical care of
neonatal animals. Literature describe specific nutrient
requirements for the infant gut and some of these nutrients are
known (e.g., glutamate, glutamine, threonine, nucleotides).
Nucleotides have been suggested to be ―conditionally
essential‖ nutrients during early weaning (Mateo et al., 2004)
because weaned pigs undergo rapid growth of tissue and organ
systems, a process that is heavily dependent on availability of
DNA, RNA, and ATP energy, whose synthesis depends on
availability of nucleotides. Neonates with intrauterine growth
retardation (IUGR) have increased morbidity and mortality early in
life and a delayed postnatal growth and development, as well as
increased susceptibility to infection (Pallotto and Kilbride, 2006).
Recently, Che et al. (2016) have shown that dietary nucleotides
supplementation improved nutrients utilization, intestinal function
and immunity of IUGR piglets. Zhang et al. (2013) have also
shown that a dietary supplementation with 220, and 275 g/ton
nucleotides improved growth performance, fecal Lactobacillus,
and blood IgG concentration in weanling pigs.
Targeted Amino-acids for their functional role could be
supplied during a certain period depending of the animal status.
For example, Le Floc’h et al. (2008) reported that inflammation
increases tryptophan (Trp) catabolism and thus may decrease
Trp availability for growth. Consequently, it may be assumed that a
provision of synthetic Trp above the requirement for growth may
increase availability for immune response and growth. Pulsatile

delivery of leucine (Leu) has been documented to be an effective
strategy to increase lean growth of neonatal piglets (Boutry et al.,
2016). Glutamic acid (Glu) and glutamine (Gln) are not usually
considered as essential nutrients; however the provision of
supplemental Gln to both suckling and weaned piglets has
demonstrated improvements in growth and health, most probably
related to improved intestinal status and immune function. L-
glutamine and glutamate has been reported to enhance intestinal-
mucosal mass and barrier function (tight junctions), and influences
the expression of amino acid receptors and transporters in the
jejunum of weaning piglets (Lin et al., 2014).
DIETARY STRATEGIES FOR PIGLETS: AFTER WEANING
Low Protein Diets. High Quality Protein Sources
High protein diets increase the luminal concentration and

epithelial exposure to putatively toxic metabolites (such as
ammonia, hydrogen sulphide, or biogenic amines) and increase
the risk for post-weaning diarrhea. Thus, the use of diets with low
levels of high-quality protein may help to reduce the risk for
intestinal disease in young pigs (Pieper et al., 2016). The dietary
CP reduction allows to reduce the N indigestible fraction and to
avoid AAs imbalance and excess.
It is well established that in feedstuffs of lower quality, more
undigested dietary proteins will enter and ferment in the hindgut. In
plant ingredients, the structural properties of proteins may play a
major role in the resistance to denaturation and gastrointestinal
digestion. For example, the β-sheet structures of raw legume
proteins and the intermolecular β-sheet aggregates, arising upon
heating were highly negatively correlated with feed digestibility
values (r =−0.980; Carbonaro et al., 2012). The decrease in
protein digestibility as dependent on β conformations can be
explained by the high hydrophobic character of these structures.
On the other hand, the presence of random coil or unordered

secondary structures in animal food proteins has been related to
an increased digestibility. Gloaguen et al. (2014) have confirmed
the possibility to formulate very low-CP (13.5%) diets that maintain
the growth of 10 to 20 kg pigs as long as AAs requirements are
satisfied by use of free AAs supplementation.
Animal plasma is a by-product from the abattoir, obtained
from animal blood. During spray drying, plasma proteins are
exposed to high temperatures for a very short period of time, and
this has the advantage over conventional drying that the proteins
are not denaturalised and preserve their biological activity. Spray-
dried animal plasma has been shown to have beneficial effects
on health status and growth performance in weaning pigs (Moretó
and Pérez-Bosque, 2009). The effects appeared to be more
efficient under a pathogen challenging environment (Coffey and
Cromwell, 1995). It has been suggested that the beneficial effects
of spray dried plasma are related to its immunoglobulin content
(Gatnau and Zimmerman, 1991). Different authors have indicated
that dietary addition of spray-dried plasma has a beneficial
influence on the health condition of weaning pigs by alleviating
liver damage, promoting intestinal development, improving
intestinal barrier function, and reducing overstimulation of immune
response (Zhang et al., 2016).
Low Calcium diets
Plant ingredients contain low Ca levels and 1/3 may be

bound to P phytate (Selle et al., 2009). Therefore, Ca has to be
supplemented with animal or mineral sources, such as limestone
or calcium phosphates. However, growth performance and feed
intake may decrease linearly with increasing dietary Ca
(González-Vega et al., 2016; Rousseau et al., 2016), and the
magnitude of the effect is exacerbated when low dietary P levels
are provided (Rousseau et al., 2016). We have also observed that
high dietary Ca level (0.95% of Ca; 1.55% of limestone) decreases
growth performance (reduces BW and ADG, and increases FCR)

as compared to 0% limestone diets (Table 2).
Table 2 - Growth performance of piglets after weaning fed on

diets containing 0 or 1.55% limestone (0.65 vs 1.1% Ca)
during the first 14 d after weaning, and a common
starter diet until d 35 post-weaning (Blavi, 2017)
Limestone 0 1.55 P- value

Dietary Ca (%) 0.35 0.9 -
BW 1 d0 (kg) 7.54 7.54 NS
BW d14 (kg) 11.30 10.63 0.009
2
ADG d0-14 (g/d) 269.2 220.7 0.008
BW d35 (kg) 22.2 21.1 0.04
1
BW= Body Weight.
2
ADG= Average Daily Gain.
3
NS: No significant.
Elevated levels of Ca may reduce phytase efficacy (Selle et

al., 2009) and P digestibility (Stein et al., 2011). Limestone also
show a high acid-binding capacity (Lawlor et al., 2005). Therefore,
high dietary Ca may also favor an increase in digestive tract pH,
which in turn decreases phytate solubility and affect protein
digestibility (Selle and Ravindran, 2008). Recently, we have
investigated whether different levels of dietary Ca in combination
with therapeutics levels of Zn modifies the gastrointestinal
microbiota and gene expression in the gastrointestinal tract of
weaned pigs (Blavi, 2017). Pigs fed high Ca levels during 14 d
after weaning showed a higher gene expression related to the
inflammatory response, and a higher heterogeneous microbial
community with increased Bacterioides genera in colon, which
may be related with the detrimental growth performance.
Palatable Ingredients
Sensory pleasure is an important determinant of behavior. It

is well known that pigs have an innate preference for sweet
(Kennedy and Baldwin, 1972) or umami taste (Tedo, 2009), and
they reject bitter, sour taste or new and unfamiliar flavors (Blair
and Fitzsimons, 1970). However, chronic stress may reduce the
ability to experience pleasure in response to typically positive
stimuli thereby displaying an analogue of anhedonia. Both social
stress (as that provoked after weaning) and restraint stress
reduced sucrose preference at low concentrations but not at
higher concentrations suggesting that weaning stress may limit
feed consumption in pigs unless a palatable feed is present
(Figueroa et al., 2015).
Selection of palatable ingredients and additives to
encourage feed intake will contribute to prevent digestive
problems related with weaning. Solà-Oriol et al. (2009a,b, 2011)
studied the preference of different feed ingredients commonly
used in piglet diets (twenty four cereals, fifteen protein sources, six
fat sources and three fiber sources) and established a ranking of
feed ingredient preferences by weanling pigs. The percentage of
preference, which was studied as a double choice for piglets as
compared to a common reference diet, was affected by the nature
of cereal, protein, fat and fiber included, and by the technological
treatments.
Feeds containing extruded rice, extruded naked oats or
naked oat were preferred to the reference diet, containing 60% of
white rice and 20% soybean meal (SBM). On the other hand, the
reference diet was preferred to the diets containing oats or cooked
oats, wheat, extruded wheat, corn, sorghum, barley, or biscuit
meal. Extrusion improved the preference of corn and naked oats,
but had no effect in barley, rice or wheat. Results showed also a
higher preference for diets presented in pelleted than in mash
form. For protein sources porcine soluble peptides (PDP) and
fishmeal at 50 g/kg and fishmeal, lupine, soybean meal-44 and
dried skimmed milk at 100 g/kg were preferred to the reference

SBM diet. On the contrary, relative to SBM an aversion was
observed for potato protein at 50 g/kg; rapeseed meal, acid milk
whey and potato protein at 100 g/kg and porcine soluble peptides,
soybean protein concentrate, wheat gluten, acid milk whey,
rapeseed meal, sunflower meal and potato protein at 200 g/kg
(Figure 4).
Figure 4 - Percentage of preference for the different protein

sources at the different inclusion levels tested. Different
letters indicated different preference values between
protein sources at each level of inclusion (P < 0.05).
Asterisks indicate that preference or aversion values
are significantly different from the neutral values of
50%. Adapted from Solà-Oriol et al. (2011).
Additives
Acidifiers
Recent studies support potential therapeutic roles for

specific fatty acids (short chain and medium chain fatty acids and
long chain polyunsaturated fatty acids) in intestinal inflammation of
pigs. In a dissociated form, organic acids reduce the pH, which
contributes to lower bacteria proliferation in the gastrointestinal
tract. Short chain fatty acids (SCFAs), for example, have been
reported to reduce gastric pH and delay the multiplication of an
enterotoxigenic E. coli. In an undissociated form, they are lipophilic
and can enter the undesirable bacteria through the cell wall. Short
chain fatty acids have many effects on the immunity response,
modulating neutrophil function and survival as well as the
cytokines they produce.
Addition of Sodium Butyrate in the diets of weaning piglets
plays an important role in recovering the intestinal tight junctions
having a positive effect on the proliferation of normal gut cells and
the maintenance of the gut integrity by enhancing mRNA
expression of the intestinal mucosal tight junction proteins. Wen et
al. (2012) reported that sodium butyrate (1 g/kg feed) improved
intestinal morphology, reduced the total viable counts of proximal
colon Clostridium and Escherichia coli, and decreased signs of
intestinal inflammation, such as TNF-α and IL-6 levels in the
serum.
Coated butyrate products are based on sodium butyrate
and calcium butyrate with a fat coating up to 70% of the total
product weight. Fat coating deals with the smell issues, provides
the protection needed to get past the acidic stomach, and makes
the product available in the small intestine. Tributyrin, like the
butyric acid present in sow milk, is an esterified form of butyrin
binding the butyrate molecule with glycerol. The butyric acid
content compared to the coated salts is much more concentrated
than the coated salts. Dietary supplementation with 0.1 %
tributyrin alleviated intestinal injury by inhibiting apoptosis,

promoting tight-junction formation (Hou et al., 2014).
Fatty acids with aliphatic tails of six to twelve carbon atoms
are called medium-chain fatty acids (MCFA, such as caproic
acid). MCFA are considered to be anionic surfactants, Besides
their acidic function, MCFA are lipophilic, they can dissolve in fats,
which, as a result of this property, have antibacterial effects (Mroz
et al., 2006). It has been shown that MCFA have a better microbial
regulatory effect, reflected in their lower minimum inhibitory
concentration values than short-chain organic acids towards a
broader spectrum of bacteria (both gram-positive and gram-
negative bacteria). That way, they can disrupt cell membranes of
pathogenic micro-organisms. In a rat model, MCFA showed a
significant decrease in the expression of proinflammatory
cytokines (TNF-α, IL-18, macrophage inflammatory protein-2 and
monocyte chemoattractant protein-1) in the ileum and Peyer’s
patches in a sepsis model of rat (Kono et al., 2010).
Phytogenics
Sensory additives are a type of feed additive defined as

―any substance, the addition of which to feed improves or changes
the organoleptic properties of the feed‖. A more recent trend in EU
feed additive innovation is the differentiation of a few botanically-
derived products as zootechnic feed additives. Phytobiotics
represent a source of various chemical and bioactive compounds,
such as terpenes, phenols, glycosides, saccharides, aldehydes,
esters and alcohols. Although the mechanisms of phytobiotics are
not entirely understood, their benefits to the overall health of
animals have been noted. Such beneficial effects claimed are
stimulation of digestive secretions; immune stimulation, intestinal
microflora modulation, anti-bacterial effects, coccidiostatic,
anthelmintic, and anti-inflammatory activities; and antioxidant
properties (Windisch et al., 2008; Wenk, 2003).
Chemical analyses of some essential oils have shown the

principal nutraceutical constituents to be Carvacrol and Thymol,
for example. In vitro, essential oil has been reported to possess
antimicrobial and antioxidant activities. Eugenol, a phytonutrient
extracted from cloves, is also a phenolic compound with previously
described functions as an antimicrobial and anti-inflammatory
agent when used at high concentrations. Eugenol acts to
strengthen the mucosal barrier by increasing the secreted Muc2
expression and thickness of the inner mucus layer, which protects
against invading pathogens and intestinal inflammation
(Wlodarska et al., 2015).
Fenugreek (Trigonella foenum graecum L.) is a legume
seed which it has been shown to affect the intestinal microbiota
and immunological responses in animals. Fenugreek had
interesting effects in piglets, lowering the pH and enterobacteria,
and increasing the concentrations of lactic acid and lactobacilli in
the digesta (Zentek et al., 2013). The literature offer also
interesting reports including among other, capsicum and turmeric
oleoresin, curcumin, limonene, etc.
After reviewing the literature, it appears evident that there is

not a unique solution to improve performance of the young pigs.
On the other hand, a complex set of dietary and management
strategies are necessary to allow an optimum and efficient growth
performance, as well as to reduce antimicrobial and therapeutic
uses of zinc oxide. In addition to the general enhancement of
animal health and welfare, with a right management of nurseries,
specific alternatives to antibiotics are necessary, including
vaccination, feeding approaches and breeding. The present report
has focused some dietary and feeding strategies to improve
performance of piglets during the first days of life and after
weaning. Strategies include a tuned feeding of sows during
gestation (to enhance fetal growth and vitality of piglets) and
lactation (to increase milk production), as well as feeding piglets

during the first days of life and after weaning with specific
nutrients, additives, palatable and digestible diets to enhance an
early feed intake, adequate microbiota, and healthy gut.
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IV INTERNATIONAL SYMPOSIUM ON

Luiz Fernando Teixeira Albino

March 29 and 30, 2017

UNIVERSIDADE FEDERAL DE VIÇOSA

March 29 and 30, 2017

Copies of this book can be purchased at:

Catalog file prepared by Catalogation Section and Classification

I61 International Symposium on Nutritional Requirement of Poultry

1. Nutrição animal - Congressos. 2. Aves – Alimentação e

III INTERNATIONAL SYMPOSIUM ON

March 29 and 30, 2017

SPECIAL THANKS TO THE COMPANIES FOR

Luiz Fernando Teixeira Albino and Horacio Santiago Rostagno

FEEDSTUFF COMPOSITION FOR POULTRY AND SWINE….. 1

NUTRITIONAL RECOMMENDATIONS FOR LAYING HENS

AMINO ACIDS ESSENTIALITY FOR SOWS: NEW APPROACHES 273

A sufficient heat-treatment of soybeans is necessary in

digestibility of Lys increased from 50.8% in T1 to 84.7% in Z2 and

Today’s livestock production is in the focus of quite different

Bellego and Noblet, 2002; Shriver et al, 2003; Lordelo et al.,

product. Excessive heat treatment may result in decreased amino

Table 1 - Processing conditions used to produce sub-batches of

FFSB - full-fat soybeans.

One batch of broken, raw soybeans was purchased from

Sub-batches of T8 were further processed at 110°C over

stability of the final product. The detailed processing conditions are

Animals and Procedures – Growth trial

The experiment was conducted at the International Poultry

Diets – Growth trial

Starter (d 1‒10), grower (d 11-24) and finisher (d 25 ‒ 35)

corresponding to 95 % of the recommended requirement. The diet

SID - standardized ileal digestible.

Animals and Procedures – Digestibility trial

The digestibility trial was conducted in cooperation with

Table 4 - Dietary composition of the assay diets

Each diet was offered ad libitum to six cages (6 birds/cage)

20 °C for chemical analyses (dry matter, titanium, nitrogen, and

Apparent ileal nitrogen and amino acid digestibility

Apparent ileal digestibility coefficient (AID, %) = ((AA / Ti) d - (AA

Apparent digestibility data were converted to standardized

Where, AID = apparent ileal digestibility coefficient of the amino acid

The raw and processed FFSB as well as commercial SBM

Association of Cereal Chemists (AOCS 1996), and trypsin inhibitor

Experimental data were analyzed as a completely

normal distribution. Hierarchical clustering of the treatments and

RESULTS AND DISCUSSION

Except for PS all other in vitro indicators responded strongly

Table 5 - In-vitro indicators of soybean processing

Although it has been suggested that the PS assay may be

soybean over-processing. As indicated by correlation coefficients

performance. Growth rate and FCR did not change appreciably at

Table 7 - Effects of soybean treatment on carcass characteristics

Table 8 - Pearson correlation coefficients describing the relations

Producing a feed that meets the nutritional requirements of

The determined coefficients of standardized ileal amino acid

Figure 1 - Hierarchical clustering of the treatments based on their

In general, the digestibility coefficients of all amino acids of

Table 11 - Comparison of the standardized ileal digestibility

SUMMARY AND CONCLUSION

Fontaine, J., U. Zimmer, P. J. Moughan, and S. M. Rutherfurd. 2007. Effect of

Leeson, S., J. O. Atteh, and J. D. Summers. 1987. Effects of increasing dietary

Ravindran, V., M. R. Abdollahi, and S. M. Bootwalla. 2014. Nutrient analysis,

The amount of energy required by poultry to maximize