1. health and fitness
2. disease
3. cholesterol

http:// www.jstage.jst.go.jp / browse / jpsa
doi:10.2141/ jpsa.0140102
Copyright Ⓒ 2015, Japan Poultry Science Association.
Effect of Supplementing Synthetic Amino Acids in Low-protein Diet and
Subsequent Re-feeding on Growth Performance, Serum Lipid Profile
and Chemical Body Composition of Broiler Chickens
Rattana Nukreaw and Chaiyapoom Bunchasak
Department of Animal Science, Faculty of Agriculture, Kasetsart University, Bangkok, Thailand
This study was conducted to evaluate the effects of supplementing methionine (Met) and lysine (Lys) in low
protein (Low-CP) diet during 1-21 days of age, and subsequent re-feeding with conventional diet during 22-42 days
of age on growth performance, serum lipid profile, chemical body composition and carcass quality of broiler chickens.
During 1-21 days of age (starter period), 480 male broiler chicks (Ross 308) were divided into three treatments and
given the following diets: 1) conventional diet group (all nutrients met the requirements of the strain), 2) Low-CP diet
without Met and Lys supplementation and 3) the Low-CP diet supplemented with Met and Lys (Low-CP + Met +
Lys). During the finisher period (22-42 days of age), all groups were fed a diet containing the same nutrients in
accordance with the recommendations of the strain. At 21 days of age, Low-CP + Met + Lys diet showed significantly better growth performance and breast meat yields than those of the Low-CP diet group. Feed and protein intake
of the chicks fed conventional diet was significantly higher than both of the other groups (P＜0.01), whereas Low-CP
+ Met + Lys diet clearly improved protein efficiency (P＜0.01). Feeding Low-CP diet increased abdominal fat
content and body energy content (P＜0.05), while the supplementing synthetic amino acids slightly decreased the fat
content. Triglyceride, very low density lipoprotein (VLDL) and T3 hormone in blood were significantly increased in
Low-CP + Met + Lys diet group compared to the conventional diet (P＜0.05). After the re-feeding phase, feeding
Low-CP diet groups were unable to compensate body weight equal to the conventional diet, although a compensation
of FCR was observed. Feeding Low-CP + Met + Lys diet showed the same breast meat yield compared to the
conventional diet, but abdominal fat, triglyceride and VLDL in blood were significantly increased (p＜0.05). In
conclusion, supplementing Met + Lys in Low-CP diet improved performance production, but was still inferior to the
conventional diet.
Key words: compensation, low-protein diet, re-feeding, synthetic amino acids
J. Poult. Sci., 52: 127-136, 2015
Introduction
Continuous genetic selection and improvement in nutrition
of broiler chickens have led to a very fast growth rate in
modern strains (Lippens et al., 2002). However, the rapid
growth rate causes several problems, namely increased body
fat deposition and some metabolic disorders such as high
incidence of skeletal diseases, sudden death syndrome as
well as ascites (Yagoub and Babiker, 2008). Excess body fat
deposition in broiler chickens is of concern to both producers
and consumers. High body fat deposition in broiler chickens
results in an inefficient of energy metabolism and overall
Received: July 4, 2014, Accepted: October 24, 2014
Released Online Advance Publication: November 25, 2014
Correspondence: Chaiyapoom Bunchasak, Department of Animal Science,
Faculty of Agriculture, Kasetsart University, Bangkok, Thailand.
(E-mail: [email protected])
feed utilization (Pasternak and Shalev, 1983), and represents
economic loss to producers (Garlich, 1979).
Quantitative feed restriction is one methodology for
limiting the amount of daily feed consumption, while
qualitative feed restriction is defined as nutrient dilution of
the diet (Leeson and Zubair, 1997). Re-feeding after quantitative feed restriction may result in improved efficiency of
growth and superior carcass characteristics of broiler
chickens (Yagoub and Babiker, 2008) therefore the early
feed restriction is applied in order to induce catch-up growth
(Susblia et al., 2003), improve efficiency of feed utilization
and reduce abdominal fat (Santoso et al., 1993). Conversely, several investigators found that quantitative feed
restriction or caloric restriction did not reduce abdominal fat
pad of broiler chickens or rats after re-feeding (Crescenzo et
al., 2010). Moreover, in some cases, phenomena of compensatory response from the re-feeding facilitate the devel-
Journal of Poultry Science, 52 (2)
128
opment of obesity (Riccardi et al., 2004) due to increased
greater lipogenic activity and the synthesized endogenous
and exogenous triglycerides obtained from the diet (Duarte et
al., 2012).
Methionine (Met) and lysine (Lys) are considered as first
and second limiting amino acids for poultry, particular in
corn-soybean diet, respectively. Met and/or Lys additions
commonly improve breast meat yield and reduce abdominal
fat content in the carcass (Rakangtong and Bunchasak,
2011). In Low-CP diets (qualitative feed restriction), Lys
and Met requirements of broilers are higher than for conventional diets for maximum weight gain and feed efficiency
(Labadan et al., 2001). Abdel-Maksoud et al. (2010)
reported that maximum body weight could be obtained with a
21% Low-CP plus amino acids supplementation which was
the same as that of the chicks fed high protein diet (23% CP).
However, inconsistencies of improvement of growth performance due to amino acid supplementation in Low-CP
diets have been observed (Keshavarz and Austic, 2004).
Recently, Nukreaw et al. (2011) suggested that reducing the
protein concentration with Met supplementation during 1-21
days of age, then re-feeding with a conventional diet is an
appropriated tool for improving overall protein utilization,
reducing fat accumulation and slightly reducing the production cost. Azarnik et al. (2010) also found that supplementing a Low-CP diet with amino acids during starter
period may partially correct the depression in the growth
Table 1.
performance of broilers after re-feeding.
Hence, it is hypothesized that complete compensatory
response may be found when feeding Low-CP diet with
amino acid supplementation and subsequent re-feeding with
conventional diet. Therefore, this study was conducted to
evaluate the effect of Met and Lys supplementation in LowCP diet during starter-grower period and followed by feeding
conventional diet during finisher period on production performance, serum lipid, chemical body composition and
carcass quality of broiler chicken at 42 days of age.
Materials and Methods
Animal and Managements
Four hundred eighty male broiler chicks (day-old, Ross
308) were used in this trial. The chicks were divided into 3
treatment groups and each group consisted of 8 replications
of 20 chicks each. The chicks were kept in floor pens (0.09
m2 floor space per bird) under an evaporative cooling house
system. The temperature was set at 32℃ at day-old and then
was decreased by 1℃ every 3 days until a final temperature
of 25℃ was reached. The lighting management and vaccinations were provided according to commercial practice.
Each pen was equipped with one hanging feeder and three
nipple drinkers. All broiler chicks were allowed access to
water and feed ad libitum throughout the experimental
period.
Composition of the experimental diets
Amount
Ingredient
Corn
Rice bran oil
Soybean meal
Full fat soybean meal
Monodicalcium phosphate
(P 21%, Ca 16%)
Lime stone
Salt
L-Lysine-HCl
DL-methionine2
L-threonine
Premix1
Sacox
Antioxidant
Corn starch
Total
Feed cost/kg (Thai Baht)
1
Conventional diet
(1-10 days)
Low-protein diet
(Low-CP)
(1-10 days)
Conventional diet
(11-21 days)
Low-protein diet
(Low-CP)
(11-21 days)
Conventional diet
(22-42 days)
54 . 876
1 . 703
36 . 194
2 . 000
2 . 313
62 . 340
1 . 001
28 . 600
2 . 000
2 . 390
(%)
55 . 399
4 . 002
34 . 172
2 . 000
2 . 042
62 . 931
3 . 277
26 . 572
2 . 000
2 . 114
60 . 24
3 . 29
25 . 43
7 . 00
1 . 91
1 . 310
0 . 210
0 . 298
0 . 409
0 . 088
0 . 500
0 . 050
0 . 050
─
1 . 330
0 . 210
─
─
0 . 213
0 . 500
0 . 050
0 . 050
1 . 33
1 . 176
0 . 215
0 . 103
0 . 293
─
0 . 500
0 . 050
0 . 050
─
1 . 196
0 . 213
─
─
0 . 118
0 . 500
0 . 050
0 . 050
0 . 98
1 . 13
0 . 22
─
0 . 19
─
0 . 50
0 . 05
0 . 05
─
100 . 00
16 . 22
100 . 00
14 . 88
100 . 00
16 . 25
100 . 00
15 . 11
100
15 . 90
Premix: vitamin A 12,000,000 IU, vitamin D3 3,000,000 IU, vitamin E 15,000 mg, vitamin K3 1,500 mg, vitamin B1 1,500 mg, vitamin B2
5,500 mg, vitamin B6 2,000 mg, vitamin B12 10 mg, nicotinic acid 25,000 mg, D-calcium pantothenate 12,000 mg, folic acid 500 mg, biotin
120 mg, biotin 120 mg, manganese 80 g, zinc 60 g, iron 40 g, copper 8 g, iodine 0.05 g, cobalt 0.10 g, selenium 0.10; filler added to 1 ton.
2
Synthetic DL-methionine was supplied by Sumitomo Chemical, Japan
Nukreaw and Bunchasak: Supplementing Synthetic Amino Acids Low-protein
Table 2.
129
Nutritional content of the experimental diets
Amount
Ingredient
Crude protein(%)
Energy (ME) kcal/kg
Ether extract (%)
Calcium (%)
Avail. Phosphorus (%)
Lysine (%)
TSAA (%)
DL-methionine(%)
Threonine (%)
Arginine (%)
Isoleucine (%)
Valine (%)
Energy:protein ratio
Conventional diet
(1-10 days)
Low-protein diet
(Low-CP)
(1-10 days)
Conventional diet
(11-21 days)
Low-protein diet
(Low-CP)
(11-21 days)
Conventional diet
(22-42 days)
22 . 00 (23 . 44)
3 , 010
4 . 67 (4 . 55)
1 . 00
0 . 50
1 . 44 (1 . 44)
1 . 09 (1 . 08)
0 . 74 (0 . 68)
0 . 93 (0 . 78)
1 . 52 (1 . 43)
0 . 98 (0 . 92)
1 . 07 (1 . 06)
136 . 82
19 . 00 (18 . 37)
3 , 010
4 . 19 (4 . 00)
1 . 01
0 . 51
1 . 02 (0 . 99)
0 . 62 (0 . 62)
0 . 30 (0 . 28)
0 . 94 (0 . 75)
1 . 30 (1 . 15)
0 . 84 (0 . 76)
0 . 93 (0 . 89)
158 . 42
21 . 00 (21 . 40)
3 , 175
6 . 94 (7 . 54)
0 . 90
0 . 45
1 . 23 (1 . 16)
0 . 95 (0 . 91)
0 . 61 (0 . 54)
0 . 81 (0 . 71)
1 . 45 (1 . 34)
0 . 94 (0 . 87)
1 . 02 (0 . 99)
151 . 19
18 . 00 (18 . 59)
3 , 175
6 . 44 (6 . 21)
0 . 91
0 . 45
0 . 96 (0 . 90)
0 . 59 (0 . 60)
0 . 29 (0 . 27)
0 . 81 (0 . 68)
1 . 22 (1 . 10)
0 . 80 (0 . 71)
0 . 88 (0 . 82)
176 . 39
19 . 00 (18 . 15)
3 , 225
7 . 26 (8 . 10)
0 . 85
0 . 42
1 . 02 (0 . 96)
0 . 80 (0 . 78)
0 . 48 (0 . 43)
0 . 73 (0 . 68)
1 . 29 (1 . 21)
0 . 84 (0 . 77)
0 . 93 (0 . 91)
169 . 74
(…) Analytical analysis value
Experimental Design and Diet
A completely randomized design was used. The broiler
chicks were fed experimental diets from 1 to 21 days of age.
Subsequently, all groups were fed a conventional diet containing 19% CP and 3,225 ME kcal/kg of energy (according
to the recommendations for the strain) from 22 to 42 days of
age. During 1-21 days of age, the 3 experimental diets
(pellet form) were provided as follows;
1) Conventional diet (22% CP during 1-10 days of age and
21% CP during 11-21 days of age, all nutrients requirements
were meet strain recommendation)
2) Low-CP diet (19% and 18% CP without DL-Met and LLys supplementation)
3) Low-CP + Met + Lys diet (19% and 18% CP with DLMet and L-Lys supplementation)
Formula of the experimental diets is shown in Table 1 and
2. Corn-soybean based diet (pellet form) was formulated
according to the recommendations of nutrients requirements
of the commercial strain, except for protein, total sulfur
amino acids (TSAA) and Lys. In order to balance amino
acids in Low-CP diet, synthetic DL-Met and L-Lys were
supplemented to meet the requirement and TSAA/Lys or
Met/Lys ratios was set as commercial recommendations.
Feed samples were collected and subsequently ground
using a 1-mm screen in a grinder. All diets were analyzed
for protein and ether extracts according to AOAC (2000)
methods. The amino acid composition of conventional and
Low-CP diets in starter and grower periods was analyzed by
Amino Acids Analyzer (AminoTac JEOL model JLC-500/V
JEOL Ltd., Tokyo, Japan).
Growth Performance
The body weight and feed intake of the chicks were
measured at 10, 21, and 42 days of age. Protein intake, protein efficiency ratio (PER), average daily gain (ADG) and
feed conversion ratio (FCR) were calculated. Mortality was
checked daily for calculation of the mortality rate.
Blood Samples and Carcass Quality
At 21 and 42 days of age, after overnight feed deprivation
(8-12 hr.), all chickens were weighed. Sixteen chickens (two
chickens per pen) were randomly selected from each
treatment group and blood samples were taken from wing
vein for determination of lipids profile in serum and hormone
in plasma. The blood samples were centrifuged at 3,000×g
for 15 minutes and the serum or plasma was decanted into
aseptically treated vials. Heparin was used in order to
prevent blood clotting in order to obtain plasma for triiodothyronine hormone (T3) determination. The serum and
plasma samples were stored at −20℃ pending analysis.
Then the chicks were sacrificed using CO2 asphyxiation
for 1.5-2.0 min. The abdominal fat (including fat surrounding the gizzard) of each bird was collected and weighed
(Cabel et al., 1987). The carcass yield of a broiler chicken
was defined as the carcass without blood, feathers and
giblets. Breast (without skin), wings, thighs and drumsticks
(with skin) were weighed and expressed as a percentage of
live weight.
Analysis of Lipid Profiles in Blood
Serum samples were analyzed for triglyceride by enzyme
colorimetric method (Test kits of Human Gesllschaft für
Biochemica und Diagnostica mbH, Co., Ltd, Wiesbaden,
Germany). Likewise, VLDL was estimated by indirect
method as serum triglyceride content was divided by five
(triglyceride/5) (Friedewald et al., 1972), while low density
lipoprotein-cholesterol (LDL-C) and high density lipoprotein-cholesterol (HDL-C) were measured by enzyme colorimetric method (Test kits of Roche Diagnostics GmbH,
Basel, Schweiz). Non-esterifies fatty acid (NEFA) was
measured by calorimetric method (Test kits of Randox
Laboratories Ltd, United Kingdom).
Journal of Poultry Science, 52 (2)
130
Triiodothyronine Hormone
Plasma samples were assayed in duplicate 25 ml aliquots
following the manufacturer’s procedures. Radioactivity was
counted for measurement of level of triiodothyronine hormone (T3) in plasma using a commercial test kit (Diagnostic
Product Corporation, Los Angeles, CA, USA). Intra- and
Inter assay CVs are 5.6 and 7.5%, respectively. Sensitivity is
6.7 ng/dl T3, as determined by the concentration at B0-2 S.D.
(n＝20).
Chemical Body Composition
For body composition analysis, at 21 and 42 days of age,
the whole body of a broiler chicken (including feathers,
abdominal fat and blood) from each replication was ground
by an industrial mincer and then homogenized by a blender
twice. Twenty gram samples of the homogenate were collected into a zip-lock bag and stored at −20℃ until chemical
analysis according to the method of AOAC (2000).
The homogenized whole body was subsequently analyzed
for moisture, crude protein, total ash and fat according to the
standard procedures of AOAC (2000). Gross energy in the
whole body was analyzed by bomb calorimeter (e2k isothermal bomb calorimeter, CAL2k, Digital Data Systems,
South Africa).
Statistical Analysis
This experiment was a completely randomized design
(CRD) with eight replications. Data were analyzed with
analysis of variance (ANOVA) procedures using the model
given below. The significance of differences between treatment group means was evaluated using Tukey’s honestly
significant difference test at a 5 and 1% probability levels.
Yij＝μ＋Ai＋εij
When; Yij is the observed response, Ai is the effect of diets
and εij is the experimental error; εij~NID (0, σ2).
Results and Discussion
Growth Performance
Effects of adding Met + Lys in Low-CP diet and subsequent re-feeding on growth performance during 1-21 and
22-42 days of age are given in Table 3.
During 1-21 days of age, adding Met + Lys in low-protein
diets (Low-CP + Met + Lys) significantly improved body
weight and FCR of broiler chicks compared to those of fed
Low-CP diet, although feeding the conventional diet group
still showed the best production performance (P＜0.01).
Feed intake and protein intake of the conventional diet group
was highest (P＜0.01). The Low-CP + Met + Lys diet group
evidently promoted PER compared to the conventional and
Low-CP diet groups (P＜0.01).
During re-feeding phase (22-42 days of age), when the
same quality of feed was given, there were no significant
differences in growth rate among treatment groups (P＞
0.05). The PER and FCR of chicks fed the Low-CP diet
were significantly better than the conventional diet group.
Nevertheless, feed and protein intakes were highest in the
conventional diet group (P＜0.05).
Poor weight gain and FCR in broilers subjected to lowprotein diets or diets with suboptimal levels of Met have been
reported (Nukreaw et al., 2011; Rakangtong and Bunchasak,
2011). Adding Met to low protein diets improved body
weight gain, PER and FCR of broiler chicks (Bunchasak et
al., 1997; Cheng et al., 1997). In the current study, feeding
a Low-CP + Met + Lys diet clearly promoted better PER than
feeding a Low-CP diet alone, although the supplementation
could not achieve the maximal growth rate and FCR see in
Effects of Met and Lys supplementation in low-protein diets and subsequent refeeding on productive performance of broiler chickens during 1-21 days of age
Table 3.
Items
During 1-21 days of age
(Low-CP phase)
Initial Body Weight (g/chick)
Final Body Weight (g/chick)
Body Weight Gain (g/chick)
Feed Intake (g/chick)
Protein Efficiency ratio (PER)
Protein intake (g/chick)
FCR
During 22-42 days of age
(Re-feeding phase)
Body Weight Gain (g/chick)
Feed Intake (g/chick)
Protein Efficiency ratio (PER)
Protein intake (g/chick)
FCR
1
Conventional Diet1
Low-protein
1
Low-CP
Low-CP + Met + Lys1
40 . 57±0 . 29
909±41 . 70A
869±41 . 68A
1210±20 . 45A
3 . 34±0 . 11B
260±4 . 39A
1 . 39±0 . 04C
40 . 56±0 . 23
758±17 . 54C
717±17 . 58C
1121±46 . 91B
3 . 46±0 . 16B
207±8 . 68B
1 . 56±0 . 07A
40 . 56±0 . 25
804±45 . 01B
764±44 . 89B
1122±53 . 68B
3 . 67±0 . 06A
207±9 . 93B
1 . 47±0 . 02B
1978±106 . 75
3778±315 . 30a
2 . 76±0 . 12a
718±59 . 91a
1 . 91±0 . 09a
1931±82 . 53
3488±67 . 88b
2 . 91±0 . 10b
663±12 . 89b
1 . 81±0 . 06b
1866±83 . 42
3467±130 . 77b
2 . 83±0 . 06ab
658±22 . 23b
1 . 86±0 . 03ab
Mean±SD
Treatment means with different superscripts in the same row are significantly different (P＜0.01).
Treatment means with different superscripts in the same row are significant different (P＜0.05).
A, B and C
a and b
Nukreaw and Bunchasak: Supplementing Synthetic Amino Acids Low-protein
131
Effects of Met and Lys supplementation in low-protein diet and subsequent refeeding on productive performance of broiler chickens during 22-42 days of age
Table 4.
Items
Final Body weight (g/chick)
Body Weight Gain (g/chick)
Feed Intake (g/chick)
Protein Efficiency Ratio
Protein intake (g/chick)
FCR
Mortality (%)
Conventional Diet1
A
2888±124 . 68
2847±124 . 76A
4988±323 . 40A
2 . 76±0 . 09B
1031±66 . 85A
1 . 75±0 . 06
1 . 87±3 . 72
Low-protein
1
Low-CP + Met + Lys1
Low-CP
B
2689±90 . 81
2649±90 . 81B
4609±81 . 85B
3 . 08±0 . 10A
860±15 . 28B
1 . 74±0 . 05
2 . 50±2 . 67
2671±106 . 76B
2630±106 . 77B
4590±173 . 64B
3 . 18±0 . 03A
826±31 . 25B
1 . 74±0 . 02
0 . 62±1 . 76
Mean±SD
A and B
Treatment means with different superscripts in the same row are significantly different (P＜0.01).
the conventional diet group. Azarnik et al. (2010) also reported that supplementation of amino acids can partially
correct the depression in growth performance observed with
Low-CP diets. Therefore, it can be said that supplementing
synthetic amino acids in low protein diet to meet the amino
acids requirement significantly improved growth performance, but is still inferior to the conventional diet.
During re-feeding phase (22-42 days of age), interestingly, it seems that body weight of chickens fed with LowCP diet without Met + Lys supplementation (amino acids
imbalance) positively responded to re-feeding more than
those fed with Low-CP + Met + Lys diet. This indicates that
the degree of compensatory growth response should be related to the amount of amino acids restriction. In pigs, a high
degree of Lys restriction has been shown to result in higher
compensatory growth response compared to those fed a
lower degree of Lys restriction (Fabian et al., 2002).
Incomplete compensation of body weight during refeeding phase may be caused by low feed consumption, since
animals usually consume higher feed in order to catch-up
their growth rate. Body weight and feed intake of chicks fed
Low-CP diets (with or without Met + Lys supplementation)
were depressed around 11-16% and 7%, respectively. Subsequently, during re-feeding phase, body weight and feed
intake of Low-CP diet groups were lower than the conventional diet group by 2-5% and 7-8%, respectively. Therefore, FCR of Low-CP diet groups were better than the conventional diet group by 2-5%. This means the suppression of
feed intake due to feeding Low-CP diet was continuously,
although chicks were fed with normal diet and FCR was
improved. The results reported by Bikker et al. (1996) indicated that a compensatory growth response mainly depends
on an increase in feed intake relative to the body weight,
whereas the relative feed consumption to body weight of
chicks fed Low-CP diet groups in the current study was
lower than the conventional diet group. Similarly, Plavnik
and Hurwitz (1988) reported that body weight of the compensatory birds did not equal that of the control group at
market age. Consequently, they reported that feed intake is
depressed by feeding diets that are severely deficient in crude
protein did not recover after realimentation (Plavnik and
Hurwitz, 1990). However, it is clear that re-feeding significantly improves FCR and PER compared to those of the
conventional diet group. This improvement may be related
to the hypertrophy of the gastrointestinal tract after feed
restriction (Rincon and Leeson, 2002).
Overall growth performance traits (1-42 days of age) are
shown in Table 4. Body weight gain, ADG, feed intake and
protein intake of the chickens fed conventional diet were
significantly higher than those of other groups (P＜0.01).
Feeding Low-CP diets (with and without Met + Lys supplementation) improved PER compared to the conventional
diet (P＜0.01). There were no significant effects of dietary
treatments on FCR or mortality rate.
For overall feeding period (1-42 days), the growth rate of
the conventional diet group was superior to the Low-CP and
Low-CP + Met + Lys diet groups due to higher feed consumption, while FCR was not significantly different, and
PER was poorer (P＜0.05). The phenomenon of chicks fed
Low-CP diets failing to catch-up their body weight may be
caused by inability to increase feed consumption. In contrast
to the growth rate, feeding Low-CP diets and subsequent refeeding clearly improved the conversion of protein intake to
body weight (PER). Zimmerman and Khajarern (1973) suggested that compensatory responses in growth performance
are not due to an increased feed consumption, but reflect a
change in metabolism. Campbell et al. (1983) and Chiba et
al. (2002) reported that pigs subjected to dietary restrictions
utilized feed more efficiently during the realimentation phase
than did unrestricted pigs. This indicates that mechanisms of
compensatory response for growth rate and efficiency of
nutrient utilization (FCR and PER) may be different.
It is known that longer periods of undernutrition cause
more difficulty to compensate weight gain (Yu and
Robinson, 1992). There are recommendations that to allow
for full body weight recovery, feed restriction should not be
longer than 7 and 5 days for male and female broilers,
respectively (Plavnik and Hurwitz, 1991). In terms of
protein restriction, Plavnik and Hurwitz (1990) showed that
ad libitum feeding of a diet containing only 9.4% CP from 8
to 14 days decreased the feed intake of broilers by some 57%.
This decrease in feed intake resulted in 41% growth
Journal of Poultry Science, 52 (2)
132
Effects of Met and Lys supplementation in low-protein diets on abdominal
fat, outer breast and inner breast of broiler chickens
Table 5.
Items
At 21 days of age
(Low-CP phase)
Abdominal fat
Outer breast
Inner breast
At 42 days of age
(After re-feeding phase)
Abdominal fat
Carcass yield
Outer breast
Inner breast
Drumstick
Thigh
Wing
Conventional Diet1
Low-protein
1
Low-CP
Low-CP + Met + Lys1
(% of body weight)
B
1 . 51±0 . 27
12 . 94±0 . 61A
3 . 35±0 . 24A
2 . 07±0 . 26A
9 . 49±0 . 63B
2 . 85±0 . 30B
1 . 82±0 . 33AB
12 . 71±0 . 92A
3 . 19±0 . 22A
(% of body weight)
b
2 . 43±0 . 43
79 . 24±0 . 98
14 . 97±0 . 84a
3 . 81±0 . 13
9 . 77±0 . 52
13 . 25±0 . 52
9 . 10±1 . 27
2 . 74±0 . 43ab
77 . 48±0 . 80
13 . 72±0 . 78b
3 . 63±0 . 13
9 . 92±0 . 15
12 . 84±0 . 61
8 . 48±1 . 22
2 . 86±0 . 34a
78 . 22±0 . 98
14 . 63±0 . 38ab
3 . 63±0 . 14
9 . 92±0 . 49
12 . 95±0 . 49
8 . 30±1 . 41
1
Mean±SD
Treatment means with different superscripts in the same row are significantly different (P＜
0.01).
a and b
Treatment means with different superscripts in the same row are significant different (P＜0.05).
A and B
retardation, which was not completely recovered after 6
weeks of realimentation (Plavnik and Hurwitz, 1990). In the
present study, the chicken fed with Low-CP diet for 21 days;
this long protein restriction period could be one of the
reasons for incomplete compensation of body weight. Lee
and Leeson (2001) also suggested that full body weight
recovery could be realized more consistently if a number of
short restriction periods were used instead of a long one.
Therefore, we can conclude that the success of a strategy of
feeding low protein diet with synthetic amino acids supplementation and consequent re-feeding conventional diet
would depend on some conditions as follows; 1) the degree
of amino acids or protein restriction, 2) duration of protein
restriction and 3) feed consumption response during refeeding phase (the reflection of metabolism)
Carcass Quality
Carcass quality of the chicks at 21 and 42 days of age are
shown in Table 5. At 21 days of age, abdominal fat of LowCP diet group was significantly higher than that of the conventional diet group (P＜0.01). Outer and inner breast meat
of the Low-CP diet group was significantly lower than both
of the other groups, while supplementation with synthetic
DL-Met and L-Lys in Low-CP diet resulted in breast meat
production and abdominal fat content equal to the conventional diet group.
After re-feeding phase, at 42 days of age, abdominal fat of
chickens fed conventional diet was significantly lower than
the Low-CP + Met + Lys diet group. Outer breast meat production of the Low-CP diet group was significantly smaller
than for the conventional diet group (P＜0.05). Differences
between the treatments with respect to carcass yield, inner
breast meat, wing, drumstick and thigh (P＞0.05).
Feeding Low-CP diet significantly decrease breast meat
yield and increase fat accumulation, while supplementation
Met + Lys prevent these negative effects at 21 days of age.
This finding is in according with generally accepted finding
that diets with low protein level increase energy retention as
fat (Swennen et al., 2004), and that supplementation with
essential amino acids in the low protein diets promotes meat
production and reduce fat content (Schutte and Pack, 1995;
Nukreaw et al., 2011). Explanations of these phenomena
have been reported extensively.
After re-feeding phase, unlike the response of feed and
protein conversion, there was no significant compensatory
response in breast meat yield. Accordingly, breast meat production of chicks fed with Low-CP diet was poorest, whilst
Low-CP-Met + Lys diet still had similar outer breast meat
compared to that of the conventional diet group. This is in
accordance with the findings of Nukreaw et al. (2011). Surprisingly, abdominal fat content in chicks fed with Low-CPMet + Lys was higher than that of the conventional diet
group, although energy consumption was not elevated and
breast meat yield production was still promoted. The mechanism for this is unknown.
Serum Lipid Profile and Hormone T3
Serum lipid profiles of broiler chickens at 21 and 42 days
of age are presented in Table 6. At 21 days of age, serum
triglyceride and very low density lipoprotein (VLDL) of the
chicks fed conventional diet group were significantly lower
than Low-CP + Met + Lys diet group (P＜0.05). Serum cholesterol, low density lipoprotein-cholesterol (LDL-C), high
density lipoprotein-cholesterol (HDL-C) and non-esterifies
fatty acid (NEFA) did not differ significantly difference
among the dietary treatments (P＞0.05). Hormone T3 was
significantly elevated when chicks were fed with Low-CP
diet, while Met + Lys supplementation in Low-CP diet sig-
Nukreaw and Bunchasak: Supplementing Synthetic Amino Acids Low-protein
133
Effects of Met and Lys supplementation in low-protein diet on serum lipid
profile and T3 of broiler chickens
Table 6.
Items
At 21 days of age
(Low-CP phase)
Triglyceride
Total Cholesterol
VLDL2
LDL-C3
HDL-C4
NEFA5
T3 (ng/dl)
At 42 days of age
(After re-feeding phase)
Triglyceride
Cholesterol
VLDL2
LDL-C3
HDL-C4
NEFA5
T3 (ng/dl)
Conventional Diet1
Low-protein
1
Low-CP
Low-CP + Met + Lys1
(mg/dl)
b
50 . 07±11 . 89
141 . 21±13 . 68
10 . 01±2 . 38b
41 . 50±9 . 54
137 . 63±19 . 54
0 . 84±0 . 15
36 . 00±9 . 92b
60 . 72±15 . 79ab
131 . 58±17 . 49
12 . 14±3 . 16ab
32 . 37±7 . 74
135 . 88±10 . 04
0 . 85±0 . 14
57 . 60±9 . 39a
91 . 83±61 . 34a
142 . 58±22 . 60
18 . 37±12 . 27a
30 . 75±6 . 09
128 . 13±34 . 66
0 . 99±0 . 23
22 . 60±5 . 36b
(mg/dl)
b
52 . 15±14 . 72
124 . 44±12 . 23
10 . 43±2 . 94b
29 . 00±8 . 37
104 . 62±9 . 65
0 . 63±2 . 94
19 . 75±8 . 17
63 . 17±16 . 62ab
117 . 66±15 . 99
12 . 63±3 . 32ab
22 . 87±2 . 29
101 . 50±17 . 54
0 . 61±3 . 32
21 . 43±19 . 85
70 . 40±23 . 55a
118 . 45±16 . 14
14 . 08±4 . 71a
29 . 87±13 . 07
94 . 75±10 . 06
0 . 67±0 . 14
25 . 71±14 . 78
1
Mean±SD
Treatment means with different superscripts in the same row are significant different (P＜0.05).
2
VLDL＝very low density lipoprotein
3
LDL-C＝low density lipoprotein-cholesterol
4
HDL-C＝high density lipoprotein-cholesterol
5
NEFA＝non-esterifies fatty acid
a and b
nificantly depressed the concentration of serum T3 hormone
(P＜0.05).
After re-feeding phase, dietary treatments did not significantly affect serum cholesterol, LDL-C, HDL-C, NEFA or
hormone T3 (P＞0.05). However, triglyceride and VLDL
concentrations of chicks fed Low-CP + Met + Lys were
significantly higher than those of the conventional diet group
(P＜0.05).
The accumulation of fat originates from plasma triglycerides which in turn derive from the diet or are synthesized in
the liver (Griffin et al., 1992). Plasma triglycerides are
detected as VLDL or LDL (Griffin et al., 1982) and, VLDLderived triglycerides are more available for fatty acid
synthesis (Griffin and Whitehead, 1982). In broilers, the
levels of VLDL and LDL are correlated to fat deposition in
the carcass. Whitehead and Griffin (1984) indicated that
plasma VLDL is a useful parameter to infer the degree of
fatness in chickens. In the present study, chicks fed the LowCP + Met + Lys diets had higher plasma triglyceride and
VLDL than those fed with conventional diet group at both 21
and 42 days of age. Likewise, Nukreaw et al. (2011) found
that adding Met to a low protein diet linearly increased the
triglyceride and VLDL concentrations in serum. The increase in serum VLDL by Met supplementation may be
caused by the stimulation of triglyceride-rich lipoprotein
secretion from the liver (Ho et al., 1989) or by depression of
the activity of lipoprotein lipase (Wegner et al., 1978). This
study also supports the hypothesis of Nukreaw et al. (2011)
that Met supplementation in young broiler chicks may
increase triglyceride transportation from the liver and depress
fat uptake from blood circulation and then, result in a high
level of serum lipid concentration and depressed abdominal
fat accumulations. Surprisingly, after the re-feeding phase,
abdominal fat was conversely increased, but feed intake and
FCR were reduced in Low-CP + Met + Lys diet group. An
explanation for these observations cannot be given.
Feeding low protein diets generally produces high
concentration of T3 in blood (Carew et al., 1997), and
plasma T3 decrease in response to restricted feed intake or
fasting (Keagy et al., 1987). In terms of protein deficiency,
Carew et al. (1997) reported that changes in circulating
levels of thyroid hormones may be a consequence of selected
amino acid deficits. Deficiency of Lys consumption seems
to have less effect on concentration of T3 hormone in blood
(Elkin et al., 1980). In contrast, Met deficiency in poultry
diets clearly elevates the level of this hormone (Bunchasak et
al., 2006; Nukreaw, 2006). In the current study, the LowCP diet group (1-21 days of age) had the significantly
highest concentration of T3, whereas supplementation with
Met + Lys in Low-CP diet depressed the T3 to the level of
the conventional diet group. The influence of low intake of
Met on thyroid function has been illustrated by Carew et al.
(2003). These authors showed that Met deficiency increases
the production or release of T3 into the blood or inhibits its
Journal of Poultry Science, 52 (2)
134
Effects of Met and Lys supplementation in low-protein diets and subsequent
re-feeding on body composition of broiler chickens
Table 7.
Items
At 21 days of age
(Low-CP phase)
Gross energy (kcal/g)
Moisture (%)
Fat (%)
Protein (%)
Total ash (%)
At 42 days of age
(After re-feeding phase)
Gross energy (kcal/g)
Moisture (%)
Fat (%)
Protein (%)
Total ash (%)
Conventional Diet1
Low-protein
1
Low-CP
Low-CP + Met + Lys1
5 . 79±0 . 27B
67 . 67±1 . 61A
15 . 17±1 . 82
17 . 80±0 . 48a
2 . 48±0 . 18
6 . 15±0 . 13A
64 . 97±0 . 77B
16 . 93±1 . 26
16 . 34±1 . 14b
2 . 67±0 . 36
6 . 02±0 . 10AB
65 . 83±1 . 37AB
16 . 43±1 . 44
17 . 70±1 . 27a
2 . 48±0 . 19
6 . 23±0 . 27
62 . 75±1 . 78
15 . 51±0 . 84
17 . 61±1 . 25
2 . 39±0 . 24
6 . 17±0 . 16
62 . 92±1 . 01
16 . 90±1 . 34
17 . 23±1 . 16
2 . 17±0 . 31
6 . 06±0 . 31
62 . 66±1 . 32
16 . 15±1 . 50
18 . 12±1 . 12
2 . 24±0 . 25
1
Mean±SD
Treatment means with different superscripts in the same row are significantly different (P＜
0.01).
a and b
Treatment means with different superscripts in the same row are significant different (P＜0.05)
A and B
normal removal compared with control chicks consuming the
same amount of feed. Therefore, it could be implied that
supplementation of the Low-CP diet with the synthetic amino
acids (Met + Lys) which resulted a reduction of plasma T3
may have been caused by Met rather than the Lys supplementation, and that the effect of deficit of dietary Met altered
plasma T3 will be dependent on the degree of deficiency
(Carew et al., 2003). However, high blood T3 concentration
in Low-CP diet group (Met and Lys deficient diet) was
normalized by re-feeding. This indicates that metabolic
disorders caused by amino acids imbalance can be returned
to the normal range.
Body Composition
The results of the analyses of whole body composition at
21 and 42 days of age are shown in Table 7. At 21 days of
age, moisture (P＜0.01) and protein (P＜0.05) content in the
Low-CP diet group was significantly lower and gross energy
content significantly higher compared with that those of
conventional diet group (P＜0.01). The dietary treatments
did not differ significantly with respect to either total fat or
ash content (P＞0.05). At 42 days of age, after the refeeding phase, dietary treatments did not significantly affect
the moisture, gross energy, fat, protein or total ash content of
the whole body of broiler chickens (P＞0.05).
The body composition of broilers is affected by many
factors such as strain, age, sex, quality and quantity of diet,
slaughter, sampling method and environmental conditions
(Koide and Ishibashi, 1995). It is clear that feeding Low-CP
without Met + Lys supplementation increased energy content
in whole body, while water and protein were significantly
decreased. Kamran et al. (2008) reported a significant decrease in the whole body protein and increase in whole body
fat content of chicks fed Low-CP diets as compared to
controls. It can be explained that feeding Low-CP diet with
amino acid imbalance results in depression of protein synthesis, with the remaining energy transferred to fat accumulation (Kamran et al., 2008). The inverse relationship
between fat and water content in whole body has already
been reported (Rosebrugh and Steele, 1985). After refeeding phase, however, there was no significant effect of
experimental diets on chemical body composition, although
body fat content was slightly high in Low-CP diet groups.
Nukreaw et al. (2011) reported that poultry fed low-protein
diets during the early rearing period had a carcass composition similar to that of control fed birds at market ages.
Since feed consumption of the Low-CP diet groups (with or
without Met + Lys supplementation) was low, the chicks
may be able to utilize body fat as energy supply to compensate their feed efficiency.
Acknowledgments
We are grateful to Sumitomo Chemical Co., Ltd., Japan
for supplying funding and the methionine and also to the staff
of the Department of Animal Science, Kasetsart University,
Thailand.
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