Directory UMM :Data Elmu:jurnal:A:Animal Feed Science and Technology:Vol87.Issue1-2.Sept2000:
Animal Feed Science and Technology
87 (2000) 85±93
The effect of DL-methionine and betaine on growth
performance and carcass characteristics in broilers$
E. Esteve-Garciaa,*, Stefan Mackb
a
Department of Animal Nutrition, Institut de Recerca i Tecnologia AgroalimentaÁries (IRTA),
Centre de Mas BoveÂ. Apartat 415, 43280 Reus, Spain
b
Degussa-HuÈls AG, Applied Technology Feed Additives, P.O. Box 13 45, D-63403 Hanau, Germany
Received 11 October 1999; received in revised form 16 February 2000; accepted 22 June 2000
Abstract
An experiment was conducted to determine if betaine could replace methionine in a methionine
de®cient diet. In order to avoid the effects of betaine as methyl group donor or as osmoprotectant or
coccidiostat enhancer, suf®cient amounts of methyl donating compounds were added and clean
conditions were used to reduce the coccidiosis challenge.
A total of 576 day-old female broiler chicks were fed one of six diets in a 3 (0, 0.6, 1.2 g/kg of
DL-methionine)2(0 and 0.5 g/kg of betaine) factorial arrangement between 0 and 41 days. The
basal diet contained 3.2 g/kg (0±21 days), 2.8 g/kg (21±38 days) and 2.5 g/kg (38±41 days), of
methionine, respectively; 3.4 g/kg (0±21 days), 3.6 g/kg (21±38 days) and 3.5 g/kg (38±41 days) of
cystine. There were eight replicates of 12 birds per treatment. Performance was determined at 21
and 41 days. At the end, carcass and breast yields were determined on nine chickens per replicate,
and abdominal fat on three chickens per replicate.
There were no signi®cant interactions between betaine and DL-methionine. DL-methionine
improved (P < 0:001) body weight and feed to gain at 21 and 41 days whereas the effects of betaine
were relatively small and not signi®cant (P > 0:10). Breast yield increased at all levels of DLmethionine addition (P < 0:001), while betaine increased carcass yield (P < 0:05). These results
suggest that betaine does not replace methionine in its function as essential amino acid in protein
metabolism, but may improve carcass yield. # 2000 Elsevier Science B.V. All rights reserved.
Keywords: Methionine; Betaine; Broiler chickens; Performance; Carcass yield
$
Parts of this manuscript were presented at the 10th European Poultry Conference, Jerusalem, Israel.
Corresponding author. Tel.: 34-977-34-32-52; fax: 34-977-34-40-55
E-mail address: enric.esteve@irta.es (E. Esteve-Garcia).
*
0377-8401/00/$ ± see front matter # 2000 Elsevier Science B.V. All rights reserved.
PII: S 0 3 7 7 - 8 4 0 1 ( 0 0 ) 0 0 1 7 4 - 7
86
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
1. Introduction
The methionine sparing effect of betaine has been the subject of some controversy.
Some experiments on maize-soyabean meal diets and several levels of coccidial
challenge have shown positive responses to betaine supplementation in methionine
de®cient diets (Virtanen and Rumsey, 1996). The responses obtained with betaine were
comparable to those of DL-methionine, when two parts of supplemental methionine were
replaced by one part of betaine. According to the authors, the diets contained choline
levels in accordance with NRC (1994) recommendations and ionophores as coccidiostats.
In contrast to these results Rostagno and Pack (1996) using a diet based on maize,
sorghum and soyabean meal and supplemental choline to provide adequate supply of
methyl groups found small and non-signi®cant responses to betaine supplementation,
which were not comparable to those found with supplemental methionine. In another
experiment, Schutte et al. (1997) fed a complex diet containing maize, wheat, tapioca,
peas, feather meal, soyabean and other minor ingredients, and a maize soy diet. All diets
were supplemented with choline. Again, the response to betaine in both diets was small
and not signi®cant, and there was no signi®cant diet by betaine interactions. It is
interesting to note that betaine signi®cantly improved oven-ready yield and breast meat
yield, although the effect on breast meat yield was inferior to that obtained with
methionine.
In pigs, Emmert et al. (1998) did not observe any response to betaine or choline in a
methionine de®cient diet. The aim of this study was to examine the effect of
supplemental levels of either DL-methionine or betaine to a methionine de®cient diet,
which was adequate in methyl donating compounds, on growth performance and carcass
quality of broiler chickens.
2. Materials and methods
A total of 576 female broiler chickens of the Ross strain were used. The feeding
program consisted of a starter diet till 21 days of age, a ®nisher diet till 38 days of age and
a withdrawal diet till 41 days of age. The ®nisher and withdrawal diets were identical,
except for the fact that the withdrawal diet did not contain halofuginone as coccidiostat
(which was used in the starter and ®nisher diets). Each treatment diet was supplemented
with 500 ppm choline (or 1 g/kg commercial choline chloride) in order to provide
adequate amounts of labile methyl donor groups. The starter diet was based on wheat,
barley, manioc, ®sh meal, full fat extruded soyabean, soyabean meal, and lard, plus a
mineral, amino acid and vitamin premix. It was estimated to contain 13.0 MJ/kg of ME,
211 g/kg protein, 12 g/kg lysine, and 6.7 g/kg methionine cystine.
The ®nisher and withdrawal diets were based on wheat, barley, full fat extruded
soyabean, meat and bone meal, lard and a premix of minerals and vitamins and amino
acids. It was estimated to contain 13.8 MJ/kg of ME, 200 g/kg protein, 11 g/kg lysine and
6.5 g/kg methionine cystine. The composition of the experimental basal diets is shown
in Table 1. The basal diets were analyzed for amino acids following the procedure of
Llames and Fontaine (1994). Results of amino acid analysis are shown in Table 2. The
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
87
Table 1
Composition of the basal diets (g/kg)
Wheat
Barley
Lard
Manioc
Full fat extruded soyabeans
Soyabean meal
Fish meal,
Meat and bone meal, 500 g CP/kg
L-lysine HCl
Calcium carbonate
Dicalcium phosphate
Salt
Choline chloride, 50%
Minerals and vitaminsa
Stenorolc
Avizyme 1200d
Starter 0±21
days
Finisher and
withdrawalb
21±41 days
385.8
100.0
32.0
80.0
159.9
189.1
20.0
504.3
40.0
30.0
1.3
7.1
17.1
2.8
1.0
4.0
0.5
1.0
315.5
69.4
13.0
1.5
7.6
11.1
2.7
1.0
4.0
0.5
1.0
Nutrient content (analysed except for metabolizable energy)
Starter
Finisher
Withdrawal
Metabolizable energy (MJ/kg)
Crude protein
Lysine
Methionine
Methionine cystine
Threonine
Arginine
Isoleucine
Leucine
Valine
Histidine
Phenylalanine
Glycine
Serine
Alanine
Aspartic acid
Glutamic acid
13.8
200
11.0
3.0
6.6
7.4
13.6
8.3
14.9
9.3
5.3
10.0
8.7
9.9
8.7
19.2
40.1
13.8
200
11.0
2.5
5.7
6.7
13.6
8.2
14.6
9.1
5.0
9.7
9.0
9.2
8.6
17.4
39.4
a
13.0
211
12.2
3.2
6.6
7.6
14.1
8.8
15.5
9.7
5.4
10.5
9.2
9.9
9.2
20.1
40.5
A sample of 1 kg of feed contains: Vitamin A: 12000 UI; Vitamin D3: 5000 UI; Vitamin E: 30 mg; Vitamin
K3: 3 mg; Vitamin B1: 2.2 mg; Vitamin B2: 8 mg; Vitamin B6: 5 mg; Vitamin B12: 11 mg; folic acid: 1.5 mg;
biotin: 150 mg; calcium pantotenate: 25 mg; nicotinic acid: 65 mg; Mn: 60 mg; Zn: 40 mg; I: 0,33 mg; Fe:
80 mg; Cu: 8 mg; Se: 0,15 mg; EtoxiquõÂn: 150 mg.
b
Withdrawal diet did not contain stenorol and was offered at 38 days.
c
Hoechst Roussel Vet GmbH, Wiesbaden, Germany.
d
Finnfeeds International Ltd., Marlborough, Wiltshire, UK.
88
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
Table 2
Analytical composition of the experimental diets
Moisture
(g/kg)
Starter
Finisher
Withdrawal
Ether
extract
(g/kg)
Crude
protein
(g/kg)
Chloride
(g/kg)
Supplemental
methionine
(g/kg)
Betaine
HCl
(g/kg)
±
0.964
T-1
T-2
T-3
T-4
T-5
T-6
109.5
109.2
104.9
105.2
106.4
105.5
77.6
75.5
74.2
76.3
75.8
75.0
205.2
207.0
206.6
203.9
205.5
207.1
4.1
4.0
3.8
4.1
4.1
4.4
0
0
0.6
0.6
1.1
1.2
T-1
T-2
T-3
T-4
T-5
T-6
104.0
100.9
101.4
98.9
99.1
104.0
98.1
99.7
96.8
101.4
98.6
96.9
203.6
201.3
201.3
195.0
202.9
203.1
3.8
4.0
3.9
4.1
4.2
4.0
0
0
0.6
0.6
1.2
1.2
T-1
T-2
T-3
T-4
T-5
T-6
103.8
105.5
100.4
103.3
104.4
104.3
98.4
97.9
94.2
98.1
98.5
99.8
201.8
202.9
203.1
203.4
201.9
201.0
3.6
3.7
3.3
3.5
3.7
3.4
0
0
0.6
0.6
1.2
1.2
1.140
1.140
1.210
1.250
1.320
1.140
1.110
1.320
different amino acid concentrations of the ®nisher and withdrawal diets may re¯ect the
composition of the different batches of ingredients used. However, it must be noted that
all diets for each period were prepared with the same batch of ingredients, and all diets
within a period had the same composition except for the supplemental betaine and
methionine. The experimental diets were analysed for supplemental methionine (NFIA,
1991) and gross chemical composition (A.O.A.C., 1984). Betaine was analyzed by ion
exchange chromatography with UV and Refractive Index detection (Johannsen,
unpublished), based on the method of RajakylaÈ and Paloposki (1983). Results are shown
in Table 2. Analyzed betaine exceeded the expected concentrations, and could be a
re¯ection of the native content of the feed ingredients which seems to be highly variable
(Kidd et al., 1997).
2.1. Experimental design
There were six experimental treatments, each consisting of eight pen replicates of 12
chickens. Table 3 further speci®es the study design.
2.2. Controls
Chicks were weighed in bulk on arrival and at 21 days. Weight of feed plus feed trough
and live weight of the birds were recorded per pen. At the end of the experiment, on day
41, chickens were leg banded for identi®cation and were individually weighed, for
89
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
Table 3
Experimental design
Treatment
MC content of starter/grower diet (g/kg)
supplementation (g/kg)
Betaine supplementation (g/kg)
Number of replicates per treatment
Number of birds per replicate
Total number of birds per treatment
T-1
T-2
T-3
T-4
T-5
T-6
0
0
0
0.5
0.6
0
0.6
0.5
1.2
0
1.2
0.5
6.6/5.7
DL-methionine
8
12
96
carcass yield and abdominal fat, or for breast yield determinations. Nine chickens per pen
were used for breast yield determination. Carcass yield included head and toes. Three
other chickens per pen were used for abdominal fat determination. Trained personnel of
the plant processed all chickens in a commercial abattoir. Specialised personnel of the
plant processed the chickens used for breast yield in the same abattoir, and deboned
breasts were obtained and weighed. Also, carcass yield was determined with the chickens
processed for breast yield at the plant. Carcass yield included head and toes. The chickens
used for abdominal fat were slaughtered, plucked and refrigerated in the processing plant,
and were stored in a cold chamber at 48C overnight. The following day, they were
eviscerated and abdominal fat was determined for each bird as a percent of live body
weight. Abdominal fat included the fat that can be manually excised from the abdominal
cavity, including that adhering to the gizzard, surrounding the bursa of Fabricius, the
cloaca, and adjacent muscles, but not the mesenteric nor the perirenal fat.
The arrangement of cages corresponds to a Randomized Complete Block Design, with
8 blocks and 6 experimental treatments. The arrangement of treatments corresponds to a
2 3 factorial with 2 levels of betaine (0 and 0.05) and 3 levels of DL-methionine (0,
0.06, and 0.12). Interactions and main effects were determined from the F values of the
ANOVA table. For data in percentages (breast yield, carcass yield, abdominal fat) the arc
sine of the square root transformation was used to stabilise the variance. Effects of
methionine level in case the F value for methionine was signi®cant at the (P < 0:05) level
were separated by Ducan's multiple range test (Duncan, 1955).
3. Results
Performance is shown in Table 4. There was a marked and highly signi®cant response
to methionine supplementation (P < 0:001) in terms of ®nal weight and feed to gain.
Body weight responded signi®cantly (P < 0:05) to all levels of methionine, but
differences in feed to gain were not signi®cant (P > 0:05) between the medium and high
level of methionine. Effect of betaine was always small and not signi®cant, although it
must be noted that it was always in the direction of improving feed to gain. Mortality also
tended to be higher at the high methionine level, but again the differences were not
signi®cant (P > 0:05).
90
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
Table 4
Body weight in grams, feed to gain ratio and liveability of chickens fed diets containing different concentrations
of DL-methionine and betainea
Dietary treatment (%)
DL-met
Age
Betaine
21 days
b
0
0
0.06
0.06
0.12
0.12
S.E.M.
0
0.05
0
0.05
0
0.05
Liveability (%)
41 days
c
BW
Fe
BW
FE
534
530
631
639
651
673
8.3
1.923
1.863
1.728
1.697
1.668
1.673
0.0210
1721
1688
1910
1929
1957
2001
31.7
2.189
2.175
2.083
2.074
2.059
2.062
0.0332
97.0
97.0
98.0
96.9
93.9
94.8
2.16
0.002
0.81
0.97
0.68
0.24
0.88
Source of variation
DL-met
Betaine
DL-met betaine
DL-met
0.001
0.20
0.27
0.001
0.11
0.31
0.001
0.70
0.47
d
0
0.06
0.12
0 vs. 0.06 and 0.12
0.06 vs. 0.12
Betained
0
0.05
532
635
662
0.01
0.01
1.893
1.713
1.681
0.01
0.06
1704
1917
1979
0.01
0.07
2.182
2.078
2.061
0.01
0.60
97.0
97.4
94.3
0.88
0.10
605
614
1.772
1.744
1862
1873
2.110
2.104
96.3
96.2
a
Body weight in kilograms per chicken.
FE cumulative feed ef®ciency to the age speci®ed.
c
Means of eight pens per treatment, 12 chickens at the start.
d
Main effect means.
b
At the end of the experiment, there was a marked and highly signi®cant response to
methionine supplementation (P < 0:001) in body weight and feed to gain. The
differences between the medium and high methionine levels were not signi®cant
(P > 0:05) but were always in the direction of improving body weight and reducing feed
to gain. The effects of betaine were small and not signi®cant (P > 0:05) although they
were in the direction of improving body weight and reducing feed to gain.
Results of the carcass measurements are shown in Table 5. There was a marked and
signi®cant response to methionine at all levels of methionine supplementation (P < 0:05)
in carcass weight, breast weight and breast yield (P < 0:05). There was also a signi®cant
effect of betaine improving carcass yield (P < 0:05) at all levels of methionine
supplementation. The effect of methionine supplementation was very pronounced on
abdominal fat (P < 0:001) but only signi®cant (P < 0:05) between the low and medium
methionine levels. Abdominal fat tended to increase with methionine supplementation
(and carcass weight) but as a percent of live body weight, there were no signi®cant
91
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
Table 5
Carcass yield, breast yield and abdominal fat of chickens fed diets containing different concentrations of
DL-methionine and betaine
DL-met
Betaine
Carcass
weight
(g)a
Carcass
yield
(%)a
Breast
weight
(g) b
Breast
yield
(%)b
Abdominal
fat (g)c
Percent
abdominal
fat (%)c
0
0
0.06
0.06
0.12
0.12
S.E.M.
P>F
DL-met
Betaine
DL-met betaine
DL-met
0
0.06
0.12
0 vs. 0.06 and 0.12
0.06 vs. 0.12
0
0.05
0
0.05
0
0.05
1385b
1365
1553
1564
1619
1643
19.55
80.3
81.3
80.9
81.6
81.1
81.3
0.26
201.7
198.5
255.9
257.3
272.4
278.4
4.68
11.6
11.7
13.3
13.4
13.6
13.8
0.16
45.3
38.0
47.3
47.8
52.0
54.9
3.65
2.50
2.26
2.49
2.43
2.71
2.67
0.16
Betaine
0
0.05
0.001
0.76
0.53
0.17
0.0165
0.38
0.001
0.72
0.64
0.001
0.45
0.99
0.0067
0.63
0.30
0.14
0.36
0.76
1376
1560
1630
0.01
0.01
80.8
81.2
81.2
0.06
0.92
200.4
256.9
275.1
0.01
0.01
11.7
13.3
13.7
0.01
0.03
42.4
47.4
51.6
0.01
0.10
2.38
2.46
2.69
0.14
0.14
1519
1527
80.8b
81.4a
243.2
245.5
12.9
13.0
47.1
46.7
2.52
2.44
a
Values are means of eight replicates of 12 chickens.
Values are means of eight replicates of nine chickens per treatment.
c
Values are means of eight replicates of three chickens per treatment.
b
treatment effects (P > 0:05). These results were also analyzed using ®nal live body
weight as a covariable (not shown). The effect of the covariable was highly signi®cant for
carcass yield and abdominal fat (P < 0:001) but not (P > 0:05) for abdominal fat as a
percent of body weight. Differences in abdominal fat due to treatments disappeared when
the covariable was introduced in the model, indicating that they were only due to body
weight. Carcass yield showed signi®cant effects due to methionine (signi®cant at
P < 0:05 between low and medium levels) and to betaine (P 0:055).
4. Discussion
The results of performance in the present experiment suggest that betaine cannot
replace methionine in a methionine de®cient diet. This is in agreement with reports from
Rostagno and Pack (1996) and Schutte et al. (1997) in the sense that the responses to
betaine are small and not signi®cant, and not comparable to those obtained with
methionine. The reason for the discrepancy between these results and those reported by
Virtanen and Rumsey (1996) is not clear since some of the studies reported by the latter
92
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
investigators were done in the presence of coccidial challenge, which reportedly enhances
the response to betaine (Matthews et al., 1995). Zimmerman et al. (1996) did not ®nd any
bene®t of betaine supplementation in the event of a coccidial challenge. It is possible that
the osmoprotective effect of betaine directly in the gut cells could help reduce the damage
caused by coccidia, and the better integrity of the gut lining could lead to more ef®cient
nutrient utilisation and improved performance.
A positive interaction between betaine and ionophores could occur due to the
impairment of choline conversion to betaine (Tyler, 1977). However, Waldenstedt et al.
(1999) found some bene®cial effect of betaine on weight gain in the occurrence of a
coccidial challenge, but the effect was independent of that of narasin. In the current
experiment and that of Schutte et al. (1997) no ionophoric coccidiostats were used;
halofuginone was employed in the former and nicarbazin during the ®rst 21 days in the
latter. It is possible that the differences between all these experiments are due to the
presence of ionophores in the diet, which could impair the conversion of choline to
betaine. Furthermore, in the present experiment and those of Rostagno and Pack (1996)
and Schutte et al. (1997) choline supplementation was rather liberal, in order to ensure
that choline was not limiting in the diet. In the experiments reported by Virtanen and
Rumsey (1996) the total level of choline and the amount of supplemental choline are not
stated, only that the diets satis®ed NRC (1994) requirements. It is possible that if the
conversion of choline to betaine was impaired by the presence of ionophores, chickens
would respond to betaine in methionine de®cient diets, and in these conditions betaine
could replace part of the methionine of the diet.
The same arguments could apply for the reported responses to betaine supplementation
in breast meat yield (Virtanen and Rosi, 1995) which were less pronounced in the
experiments of Rostagno and Pack (1996), those of Schutte et al. (1997) and in the
current experiment. On the other hand, it is interesting to note a consistent effect of
betaine in improving carcass yield in all experiments. In the present experiment, carcass
yield was determined in the processing plant with mechanical evisceration, which only
removes intestines and part of the abdominal fat. These results would suggest that the
effect of betaine could be caused by a reduction in the weight of intestines, but in view of
the osmotic effects of betaine, it could be also due to increased water retention. Recently,
McDevitt et al. (1999) found a signi®cant reduction in relative visceral mass by betaine
and DL-methionine, which suggests that the effect of betaine observed in the current
experiment can be due to a reduction of the relative weight of the intestines. It is
interesting to note that in the current experiment DL-methionine also increased carcass
yield, although the effect was not signi®cant at the P < 0:05 level. The results of
McDevitt et al. (1999) suggest that this effect of DL-methionine on promoting carcass
yield could be real.
5. Conclusion
It can be concluded that in the conditions of the experiment, that is, in the absence of a
coccidial challenge (chickens were raised on wire cages) and in the presence of
halofuginone (stenorol) as coccidiostat, betaine does not replace methionine to a
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
93
signi®cant extend, in terms of performance and breast yield. However, betaine
signi®cantly improved carcass yield.
Acknowledgements
The technical assistance of Mr. Lluis Llaurado is greatly appreciated.
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Res. 5, 150±154.
Schutte, J.B., de Jong, J., Smink, W., Pack, M., 1997. Replacement value for DL-methionine in male broiler
chicks. Poult. Sci. 76, 321±325.
Tyler, D.D., 1977. Transport and oxidation of choline by liver mitochondria. Biochem. J. 166, 571±581.
Virtanen, E.I., Rosi, L., 1995. Effects of betaine on methionine requirement of broilers under various
environmental conditions. In: Proceedings of the 8th Australian Poultry Science Symposium, Adelaide,
pp. 88±92.
Virtanen, E., Rumsey, G., 1996. Betaine supplementation can optimize use of methionine, choline in diets.
Feedstuffs 68 (42), 12±13.
Waldenstedt, L., Elwinger, K., Thebo, P., Uggla, A., 1999. Effect of betaine supplement on broiler performance
during an experimental coccidial infection. Poult. Sci. 78, 182±189.
Zimmerman, N.G., Twining, P., Hater-Dennis, J., 1996. Betaine as a methionine substitute and coccidial
deterrent in broilers. Poult. Sci. 75, 154.
87 (2000) 85±93
The effect of DL-methionine and betaine on growth
performance and carcass characteristics in broilers$
E. Esteve-Garciaa,*, Stefan Mackb
a
Department of Animal Nutrition, Institut de Recerca i Tecnologia AgroalimentaÁries (IRTA),
Centre de Mas BoveÂ. Apartat 415, 43280 Reus, Spain
b
Degussa-HuÈls AG, Applied Technology Feed Additives, P.O. Box 13 45, D-63403 Hanau, Germany
Received 11 October 1999; received in revised form 16 February 2000; accepted 22 June 2000
Abstract
An experiment was conducted to determine if betaine could replace methionine in a methionine
de®cient diet. In order to avoid the effects of betaine as methyl group donor or as osmoprotectant or
coccidiostat enhancer, suf®cient amounts of methyl donating compounds were added and clean
conditions were used to reduce the coccidiosis challenge.
A total of 576 day-old female broiler chicks were fed one of six diets in a 3 (0, 0.6, 1.2 g/kg of
DL-methionine)2(0 and 0.5 g/kg of betaine) factorial arrangement between 0 and 41 days. The
basal diet contained 3.2 g/kg (0±21 days), 2.8 g/kg (21±38 days) and 2.5 g/kg (38±41 days), of
methionine, respectively; 3.4 g/kg (0±21 days), 3.6 g/kg (21±38 days) and 3.5 g/kg (38±41 days) of
cystine. There were eight replicates of 12 birds per treatment. Performance was determined at 21
and 41 days. At the end, carcass and breast yields were determined on nine chickens per replicate,
and abdominal fat on three chickens per replicate.
There were no signi®cant interactions between betaine and DL-methionine. DL-methionine
improved (P < 0:001) body weight and feed to gain at 21 and 41 days whereas the effects of betaine
were relatively small and not signi®cant (P > 0:10). Breast yield increased at all levels of DLmethionine addition (P < 0:001), while betaine increased carcass yield (P < 0:05). These results
suggest that betaine does not replace methionine in its function as essential amino acid in protein
metabolism, but may improve carcass yield. # 2000 Elsevier Science B.V. All rights reserved.
Keywords: Methionine; Betaine; Broiler chickens; Performance; Carcass yield
$
Parts of this manuscript were presented at the 10th European Poultry Conference, Jerusalem, Israel.
Corresponding author. Tel.: 34-977-34-32-52; fax: 34-977-34-40-55
E-mail address: enric.esteve@irta.es (E. Esteve-Garcia).
*
0377-8401/00/$ ± see front matter # 2000 Elsevier Science B.V. All rights reserved.
PII: S 0 3 7 7 - 8 4 0 1 ( 0 0 ) 0 0 1 7 4 - 7
86
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
1. Introduction
The methionine sparing effect of betaine has been the subject of some controversy.
Some experiments on maize-soyabean meal diets and several levels of coccidial
challenge have shown positive responses to betaine supplementation in methionine
de®cient diets (Virtanen and Rumsey, 1996). The responses obtained with betaine were
comparable to those of DL-methionine, when two parts of supplemental methionine were
replaced by one part of betaine. According to the authors, the diets contained choline
levels in accordance with NRC (1994) recommendations and ionophores as coccidiostats.
In contrast to these results Rostagno and Pack (1996) using a diet based on maize,
sorghum and soyabean meal and supplemental choline to provide adequate supply of
methyl groups found small and non-signi®cant responses to betaine supplementation,
which were not comparable to those found with supplemental methionine. In another
experiment, Schutte et al. (1997) fed a complex diet containing maize, wheat, tapioca,
peas, feather meal, soyabean and other minor ingredients, and a maize soy diet. All diets
were supplemented with choline. Again, the response to betaine in both diets was small
and not signi®cant, and there was no signi®cant diet by betaine interactions. It is
interesting to note that betaine signi®cantly improved oven-ready yield and breast meat
yield, although the effect on breast meat yield was inferior to that obtained with
methionine.
In pigs, Emmert et al. (1998) did not observe any response to betaine or choline in a
methionine de®cient diet. The aim of this study was to examine the effect of
supplemental levels of either DL-methionine or betaine to a methionine de®cient diet,
which was adequate in methyl donating compounds, on growth performance and carcass
quality of broiler chickens.
2. Materials and methods
A total of 576 female broiler chickens of the Ross strain were used. The feeding
program consisted of a starter diet till 21 days of age, a ®nisher diet till 38 days of age and
a withdrawal diet till 41 days of age. The ®nisher and withdrawal diets were identical,
except for the fact that the withdrawal diet did not contain halofuginone as coccidiostat
(which was used in the starter and ®nisher diets). Each treatment diet was supplemented
with 500 ppm choline (or 1 g/kg commercial choline chloride) in order to provide
adequate amounts of labile methyl donor groups. The starter diet was based on wheat,
barley, manioc, ®sh meal, full fat extruded soyabean, soyabean meal, and lard, plus a
mineral, amino acid and vitamin premix. It was estimated to contain 13.0 MJ/kg of ME,
211 g/kg protein, 12 g/kg lysine, and 6.7 g/kg methionine cystine.
The ®nisher and withdrawal diets were based on wheat, barley, full fat extruded
soyabean, meat and bone meal, lard and a premix of minerals and vitamins and amino
acids. It was estimated to contain 13.8 MJ/kg of ME, 200 g/kg protein, 11 g/kg lysine and
6.5 g/kg methionine cystine. The composition of the experimental basal diets is shown
in Table 1. The basal diets were analyzed for amino acids following the procedure of
Llames and Fontaine (1994). Results of amino acid analysis are shown in Table 2. The
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
87
Table 1
Composition of the basal diets (g/kg)
Wheat
Barley
Lard
Manioc
Full fat extruded soyabeans
Soyabean meal
Fish meal,
Meat and bone meal, 500 g CP/kg
L-lysine HCl
Calcium carbonate
Dicalcium phosphate
Salt
Choline chloride, 50%
Minerals and vitaminsa
Stenorolc
Avizyme 1200d
Starter 0±21
days
Finisher and
withdrawalb
21±41 days
385.8
100.0
32.0
80.0
159.9
189.1
20.0
504.3
40.0
30.0
1.3
7.1
17.1
2.8
1.0
4.0
0.5
1.0
315.5
69.4
13.0
1.5
7.6
11.1
2.7
1.0
4.0
0.5
1.0
Nutrient content (analysed except for metabolizable energy)
Starter
Finisher
Withdrawal
Metabolizable energy (MJ/kg)
Crude protein
Lysine
Methionine
Methionine cystine
Threonine
Arginine
Isoleucine
Leucine
Valine
Histidine
Phenylalanine
Glycine
Serine
Alanine
Aspartic acid
Glutamic acid
13.8
200
11.0
3.0
6.6
7.4
13.6
8.3
14.9
9.3
5.3
10.0
8.7
9.9
8.7
19.2
40.1
13.8
200
11.0
2.5
5.7
6.7
13.6
8.2
14.6
9.1
5.0
9.7
9.0
9.2
8.6
17.4
39.4
a
13.0
211
12.2
3.2
6.6
7.6
14.1
8.8
15.5
9.7
5.4
10.5
9.2
9.9
9.2
20.1
40.5
A sample of 1 kg of feed contains: Vitamin A: 12000 UI; Vitamin D3: 5000 UI; Vitamin E: 30 mg; Vitamin
K3: 3 mg; Vitamin B1: 2.2 mg; Vitamin B2: 8 mg; Vitamin B6: 5 mg; Vitamin B12: 11 mg; folic acid: 1.5 mg;
biotin: 150 mg; calcium pantotenate: 25 mg; nicotinic acid: 65 mg; Mn: 60 mg; Zn: 40 mg; I: 0,33 mg; Fe:
80 mg; Cu: 8 mg; Se: 0,15 mg; EtoxiquõÂn: 150 mg.
b
Withdrawal diet did not contain stenorol and was offered at 38 days.
c
Hoechst Roussel Vet GmbH, Wiesbaden, Germany.
d
Finnfeeds International Ltd., Marlborough, Wiltshire, UK.
88
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
Table 2
Analytical composition of the experimental diets
Moisture
(g/kg)
Starter
Finisher
Withdrawal
Ether
extract
(g/kg)
Crude
protein
(g/kg)
Chloride
(g/kg)
Supplemental
methionine
(g/kg)
Betaine
HCl
(g/kg)
±
0.964
T-1
T-2
T-3
T-4
T-5
T-6
109.5
109.2
104.9
105.2
106.4
105.5
77.6
75.5
74.2
76.3
75.8
75.0
205.2
207.0
206.6
203.9
205.5
207.1
4.1
4.0
3.8
4.1
4.1
4.4
0
0
0.6
0.6
1.1
1.2
T-1
T-2
T-3
T-4
T-5
T-6
104.0
100.9
101.4
98.9
99.1
104.0
98.1
99.7
96.8
101.4
98.6
96.9
203.6
201.3
201.3
195.0
202.9
203.1
3.8
4.0
3.9
4.1
4.2
4.0
0
0
0.6
0.6
1.2
1.2
T-1
T-2
T-3
T-4
T-5
T-6
103.8
105.5
100.4
103.3
104.4
104.3
98.4
97.9
94.2
98.1
98.5
99.8
201.8
202.9
203.1
203.4
201.9
201.0
3.6
3.7
3.3
3.5
3.7
3.4
0
0
0.6
0.6
1.2
1.2
1.140
1.140
1.210
1.250
1.320
1.140
1.110
1.320
different amino acid concentrations of the ®nisher and withdrawal diets may re¯ect the
composition of the different batches of ingredients used. However, it must be noted that
all diets for each period were prepared with the same batch of ingredients, and all diets
within a period had the same composition except for the supplemental betaine and
methionine. The experimental diets were analysed for supplemental methionine (NFIA,
1991) and gross chemical composition (A.O.A.C., 1984). Betaine was analyzed by ion
exchange chromatography with UV and Refractive Index detection (Johannsen,
unpublished), based on the method of RajakylaÈ and Paloposki (1983). Results are shown
in Table 2. Analyzed betaine exceeded the expected concentrations, and could be a
re¯ection of the native content of the feed ingredients which seems to be highly variable
(Kidd et al., 1997).
2.1. Experimental design
There were six experimental treatments, each consisting of eight pen replicates of 12
chickens. Table 3 further speci®es the study design.
2.2. Controls
Chicks were weighed in bulk on arrival and at 21 days. Weight of feed plus feed trough
and live weight of the birds were recorded per pen. At the end of the experiment, on day
41, chickens were leg banded for identi®cation and were individually weighed, for
89
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
Table 3
Experimental design
Treatment
MC content of starter/grower diet (g/kg)
supplementation (g/kg)
Betaine supplementation (g/kg)
Number of replicates per treatment
Number of birds per replicate
Total number of birds per treatment
T-1
T-2
T-3
T-4
T-5
T-6
0
0
0
0.5
0.6
0
0.6
0.5
1.2
0
1.2
0.5
6.6/5.7
DL-methionine
8
12
96
carcass yield and abdominal fat, or for breast yield determinations. Nine chickens per pen
were used for breast yield determination. Carcass yield included head and toes. Three
other chickens per pen were used for abdominal fat determination. Trained personnel of
the plant processed all chickens in a commercial abattoir. Specialised personnel of the
plant processed the chickens used for breast yield in the same abattoir, and deboned
breasts were obtained and weighed. Also, carcass yield was determined with the chickens
processed for breast yield at the plant. Carcass yield included head and toes. The chickens
used for abdominal fat were slaughtered, plucked and refrigerated in the processing plant,
and were stored in a cold chamber at 48C overnight. The following day, they were
eviscerated and abdominal fat was determined for each bird as a percent of live body
weight. Abdominal fat included the fat that can be manually excised from the abdominal
cavity, including that adhering to the gizzard, surrounding the bursa of Fabricius, the
cloaca, and adjacent muscles, but not the mesenteric nor the perirenal fat.
The arrangement of cages corresponds to a Randomized Complete Block Design, with
8 blocks and 6 experimental treatments. The arrangement of treatments corresponds to a
2 3 factorial with 2 levels of betaine (0 and 0.05) and 3 levels of DL-methionine (0,
0.06, and 0.12). Interactions and main effects were determined from the F values of the
ANOVA table. For data in percentages (breast yield, carcass yield, abdominal fat) the arc
sine of the square root transformation was used to stabilise the variance. Effects of
methionine level in case the F value for methionine was signi®cant at the (P < 0:05) level
were separated by Ducan's multiple range test (Duncan, 1955).
3. Results
Performance is shown in Table 4. There was a marked and highly signi®cant response
to methionine supplementation (P < 0:001) in terms of ®nal weight and feed to gain.
Body weight responded signi®cantly (P < 0:05) to all levels of methionine, but
differences in feed to gain were not signi®cant (P > 0:05) between the medium and high
level of methionine. Effect of betaine was always small and not signi®cant, although it
must be noted that it was always in the direction of improving feed to gain. Mortality also
tended to be higher at the high methionine level, but again the differences were not
signi®cant (P > 0:05).
90
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
Table 4
Body weight in grams, feed to gain ratio and liveability of chickens fed diets containing different concentrations
of DL-methionine and betainea
Dietary treatment (%)
DL-met
Age
Betaine
21 days
b
0
0
0.06
0.06
0.12
0.12
S.E.M.
0
0.05
0
0.05
0
0.05
Liveability (%)
41 days
c
BW
Fe
BW
FE
534
530
631
639
651
673
8.3
1.923
1.863
1.728
1.697
1.668
1.673
0.0210
1721
1688
1910
1929
1957
2001
31.7
2.189
2.175
2.083
2.074
2.059
2.062
0.0332
97.0
97.0
98.0
96.9
93.9
94.8
2.16
0.002
0.81
0.97
0.68
0.24
0.88
Source of variation
DL-met
Betaine
DL-met betaine
DL-met
0.001
0.20
0.27
0.001
0.11
0.31
0.001
0.70
0.47
d
0
0.06
0.12
0 vs. 0.06 and 0.12
0.06 vs. 0.12
Betained
0
0.05
532
635
662
0.01
0.01
1.893
1.713
1.681
0.01
0.06
1704
1917
1979
0.01
0.07
2.182
2.078
2.061
0.01
0.60
97.0
97.4
94.3
0.88
0.10
605
614
1.772
1.744
1862
1873
2.110
2.104
96.3
96.2
a
Body weight in kilograms per chicken.
FE cumulative feed ef®ciency to the age speci®ed.
c
Means of eight pens per treatment, 12 chickens at the start.
d
Main effect means.
b
At the end of the experiment, there was a marked and highly signi®cant response to
methionine supplementation (P < 0:001) in body weight and feed to gain. The
differences between the medium and high methionine levels were not signi®cant
(P > 0:05) but were always in the direction of improving body weight and reducing feed
to gain. The effects of betaine were small and not signi®cant (P > 0:05) although they
were in the direction of improving body weight and reducing feed to gain.
Results of the carcass measurements are shown in Table 5. There was a marked and
signi®cant response to methionine at all levels of methionine supplementation (P < 0:05)
in carcass weight, breast weight and breast yield (P < 0:05). There was also a signi®cant
effect of betaine improving carcass yield (P < 0:05) at all levels of methionine
supplementation. The effect of methionine supplementation was very pronounced on
abdominal fat (P < 0:001) but only signi®cant (P < 0:05) between the low and medium
methionine levels. Abdominal fat tended to increase with methionine supplementation
(and carcass weight) but as a percent of live body weight, there were no signi®cant
91
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
Table 5
Carcass yield, breast yield and abdominal fat of chickens fed diets containing different concentrations of
DL-methionine and betaine
DL-met
Betaine
Carcass
weight
(g)a
Carcass
yield
(%)a
Breast
weight
(g) b
Breast
yield
(%)b
Abdominal
fat (g)c
Percent
abdominal
fat (%)c
0
0
0.06
0.06
0.12
0.12
S.E.M.
P>F
DL-met
Betaine
DL-met betaine
DL-met
0
0.06
0.12
0 vs. 0.06 and 0.12
0.06 vs. 0.12
0
0.05
0
0.05
0
0.05
1385b
1365
1553
1564
1619
1643
19.55
80.3
81.3
80.9
81.6
81.1
81.3
0.26
201.7
198.5
255.9
257.3
272.4
278.4
4.68
11.6
11.7
13.3
13.4
13.6
13.8
0.16
45.3
38.0
47.3
47.8
52.0
54.9
3.65
2.50
2.26
2.49
2.43
2.71
2.67
0.16
Betaine
0
0.05
0.001
0.76
0.53
0.17
0.0165
0.38
0.001
0.72
0.64
0.001
0.45
0.99
0.0067
0.63
0.30
0.14
0.36
0.76
1376
1560
1630
0.01
0.01
80.8
81.2
81.2
0.06
0.92
200.4
256.9
275.1
0.01
0.01
11.7
13.3
13.7
0.01
0.03
42.4
47.4
51.6
0.01
0.10
2.38
2.46
2.69
0.14
0.14
1519
1527
80.8b
81.4a
243.2
245.5
12.9
13.0
47.1
46.7
2.52
2.44
a
Values are means of eight replicates of 12 chickens.
Values are means of eight replicates of nine chickens per treatment.
c
Values are means of eight replicates of three chickens per treatment.
b
treatment effects (P > 0:05). These results were also analyzed using ®nal live body
weight as a covariable (not shown). The effect of the covariable was highly signi®cant for
carcass yield and abdominal fat (P < 0:001) but not (P > 0:05) for abdominal fat as a
percent of body weight. Differences in abdominal fat due to treatments disappeared when
the covariable was introduced in the model, indicating that they were only due to body
weight. Carcass yield showed signi®cant effects due to methionine (signi®cant at
P < 0:05 between low and medium levels) and to betaine (P 0:055).
4. Discussion
The results of performance in the present experiment suggest that betaine cannot
replace methionine in a methionine de®cient diet. This is in agreement with reports from
Rostagno and Pack (1996) and Schutte et al. (1997) in the sense that the responses to
betaine are small and not signi®cant, and not comparable to those obtained with
methionine. The reason for the discrepancy between these results and those reported by
Virtanen and Rumsey (1996) is not clear since some of the studies reported by the latter
92
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
investigators were done in the presence of coccidial challenge, which reportedly enhances
the response to betaine (Matthews et al., 1995). Zimmerman et al. (1996) did not ®nd any
bene®t of betaine supplementation in the event of a coccidial challenge. It is possible that
the osmoprotective effect of betaine directly in the gut cells could help reduce the damage
caused by coccidia, and the better integrity of the gut lining could lead to more ef®cient
nutrient utilisation and improved performance.
A positive interaction between betaine and ionophores could occur due to the
impairment of choline conversion to betaine (Tyler, 1977). However, Waldenstedt et al.
(1999) found some bene®cial effect of betaine on weight gain in the occurrence of a
coccidial challenge, but the effect was independent of that of narasin. In the current
experiment and that of Schutte et al. (1997) no ionophoric coccidiostats were used;
halofuginone was employed in the former and nicarbazin during the ®rst 21 days in the
latter. It is possible that the differences between all these experiments are due to the
presence of ionophores in the diet, which could impair the conversion of choline to
betaine. Furthermore, in the present experiment and those of Rostagno and Pack (1996)
and Schutte et al. (1997) choline supplementation was rather liberal, in order to ensure
that choline was not limiting in the diet. In the experiments reported by Virtanen and
Rumsey (1996) the total level of choline and the amount of supplemental choline are not
stated, only that the diets satis®ed NRC (1994) requirements. It is possible that if the
conversion of choline to betaine was impaired by the presence of ionophores, chickens
would respond to betaine in methionine de®cient diets, and in these conditions betaine
could replace part of the methionine of the diet.
The same arguments could apply for the reported responses to betaine supplementation
in breast meat yield (Virtanen and Rosi, 1995) which were less pronounced in the
experiments of Rostagno and Pack (1996), those of Schutte et al. (1997) and in the
current experiment. On the other hand, it is interesting to note a consistent effect of
betaine in improving carcass yield in all experiments. In the present experiment, carcass
yield was determined in the processing plant with mechanical evisceration, which only
removes intestines and part of the abdominal fat. These results would suggest that the
effect of betaine could be caused by a reduction in the weight of intestines, but in view of
the osmotic effects of betaine, it could be also due to increased water retention. Recently,
McDevitt et al. (1999) found a signi®cant reduction in relative visceral mass by betaine
and DL-methionine, which suggests that the effect of betaine observed in the current
experiment can be due to a reduction of the relative weight of the intestines. It is
interesting to note that in the current experiment DL-methionine also increased carcass
yield, although the effect was not signi®cant at the P < 0:05 level. The results of
McDevitt et al. (1999) suggest that this effect of DL-methionine on promoting carcass
yield could be real.
5. Conclusion
It can be concluded that in the conditions of the experiment, that is, in the absence of a
coccidial challenge (chickens were raised on wire cages) and in the presence of
halofuginone (stenorol) as coccidiostat, betaine does not replace methionine to a
E. Esteve-Garcia, S. Mack / Animal Feed Science and Technology 87 (2000) 85±93
93
signi®cant extend, in terms of performance and breast yield. However, betaine
signi®cantly improved carcass yield.
Acknowledgements
The technical assistance of Mr. Lluis Llaurado is greatly appreciated.
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