©2007 Poultry Science Association, Inc.

Effects of Different Forced Molting Methods

on Postmolt Production, Corticosterone Level,
and Immune Response to Sheep
Red Blood Cells in Laying Hens

E. E. Onbaşılar*1 and H. Erol†

*Ankara University, Faculty of Veterinary Medicine, Department of Animal Science,

06110, Ankara, Turkey; and †Abant İzzet Baysal University, Mudurnu Süreyya
Astarcı Vocational School of Higher Education, 14800, Bolu, Turkey

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Primary Audience: Poultry Extension Specialists, Egg Producers, Researchers

SUMMARY
The objective of this study was to evaluate the effects of different molting methods on postmolt
production, plasma corticosterone levels, and antibody production to SRBC for the welfare of
laying hens. This experiment was conducted with 120 IGH-type Brown laying hens (70 wk of age),
randomly divided into 3 experimental groups. The hens in one group were fed a whole-grain barley
diet during the first 10 d (WB diet). On d 11, hens consumed 100 g of layer diet/d until d 28. In
the second group, hens were fed a Zn diet containing 10,000 mg/kg of Zn as ZnO for 10 d (Zn
diet). Hens were then provided 100 g of a layer diet from d 11 to 28. In the third group, feed was
withdrawn for 10 d, and on d 11 hens were fed a cracked corn diet ad libitum until d 28 (California
method; CAL diet). Hens in all groups were returned to the layer diet ad libitum on d 29. Egg
weight was lower in the Zn treatment than in the other treatments. Feed intake and plasma
corticosterone levels were higher and antibody production was lower in the CAL treatment than
in the WB and Zn groups. The FCR was better in the WB than in the Zn group. Mortality, egg
production, and egg quality were not significantly different among the molting methods. As a
result, the WB molting program was the best method for postmolt production among the
programs examined.

Key words: molting, laying hen, postmolt production, corticosterone level, immune response
2007 J. Appl. Poult. Res. 16:529–536
doi:10.3382/japr.2006-00089

DESCRIPTION OF PROBLEM Most researchers have reported that induced

In the commercial egg industry, widely dif-
ferent molt techniques are used for extending
laying flock performance. Molting programs are
designed to prolong the productive life of layers
and breeders. This will depend on local eco-
nomic conditions and, in particular, egg prices.
molting improves the postmolt performance
(such as egg production, egg quality, and egg
weight) of laying hens compared with the pre-
molt performance [1, 2].
Conventional induced molting methods have
used fasting of hens for shorter periods or to a
targeted BW with or without water restriction

1
Corresponding author: obasilar@veterinary.ankara.edu.tr
530 JAPR: Research Report

Table 1. Description of WB-, Zn-, and CAL-induced molting programs1

Molting Day of Light,

program experiment Feed Water h/d

WB 1 to 10 Ad libitum whole-grain barley diet Provided 8

11 to 28 100 g of layer diet per hen Provided 8
29 Ad libitum layer diet Provided 16
Zn 1 to 10 Ad libitum Zn diet containing 10,000 mg/kg of Zn as ZnO Provided 8
11 to 28 100 g of layer diet per hen Provided 8
29 Ad libitum layer diet Provided 16
CAL 1 to 10 None Provided 8
11 to 28 Ad libitum cracked corn diet Provided 8
29 Ad libitum layer diet Provided 16
1
WB = whole-grain barley method; Zn = high dietary Zn method (containing 10,000 mg/kg of Zn as ZnO); CAL = California
method.

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[2, 3, 4]. This method efficiently induces a molt
because it is management friendly and economi-
cally advantageous, and it results in satisfactory
postmolt performance for the commercial layer
industry [5]. But, recent concerns have been
raised about animal welfare during the fasting
period, because it is thought to be harmful to
the hens [6]. The other methods are feeding high
dietary Zn [7, 8, 9, 10, 11] or iodine [12], feeding
low-Ca [4, 9] or low-Na diets [13], and feeding
high-corn diets [14, 15]. The aim of the current
study was to detect the effects of different molt-
ing procedures on the postmolt production, phys-
iology, and immune response to SRBC in lay-
ing hens.
MATERIALS AND METHODS
Animals and Diets
A total of 120 IGH-type Brown laying hens
[16], 70 wk of age, were used in the present
study. The hens were housed in individual cages.

Table 2. Ingredients and chemical composition of diets

Layer Zn Cracked Whole-grain

Item diet diet corn diet barley diet

Ingredient, %
Corn 52.00 56.36 100 —
Barley 13.40 7.00 — 100
Soybean meal 21.00 21.80 — —
Fish meal 3.00 3.00 — —
Limestone 9.00 9.00 — —
Dicalcium phosphate 1.00 1.00 — —
Salt 0.25 0.25 — —
Vitamin-mineral premix1 0.25 0.25 — —
DL-Met 0.10 0.10 — —
ZnO — 1.24 — —
Chemical composition (analyzed)
ME,2 kcal/kg 2,637 2,628 3,365 2,670
CP, % 16.30 16.39 8.00 10.90
Ca, % 3.10 3.18 0.04 0.22
P (total), % 0.68 0.72 0.04 0.35
Zn,3 mg/kg 84 10,084 18 22
1
Composition per 2.5 kg: 12,000,000 IU of vitamin A, 2,400,000 IU of vitamin D3, 30 g of vitamin E, 2.5 g of vitamin K3,
2.5 g of vitamin B1, 6 g of vitamin B2, 4 g of vitamin B6, 20 mg of vitamin B12, 25 g of niacin, 8 g of calcium D-panthotenate,
1 g of folic acid, 50 g of vitamin C, 50 mg of D-biotin, 150 g of choline chloride, 1.5 g of canthaxanthin, 0.5 g of apo-
carotenoic acid ester, 80 g of Mn, 60 g of Zn, 60 g of Fe, 5 g of Cu, 1 g of I, 0.5 g of Co, and 0.15 g of Se.
2
The ME content of diets was estimated by using a prediction equation [24].
3
Calculated based on the feed composition tables of the NRC [37].
ONBAŞıLAR AND EROL: MOLTING METHODS AND PERFORMANCE 531

Table 3. Effect of different induced molting programs on BW of brown laying hens

Molting program1

WB Zn CAL
Time of
experiment n Mean ± SEM n Mean ± SEM n Mean ± SEM P-value

Initial BW, g 40 2,122 ± 55.56 40 2,173 ± 34.05 40 2,204 ± 53.26 NS

10-d BW, g 40 1,746 ± 48.69b 40 2,011 ± 37.17a 40 1,768 ± 43.86b <0.001
28-d BW, g 35 2,003 ± 37.93a 38 2,012 ± 34.31a 32 1,859 ± 53.37b 0.021
10-d BW loss, % 40 18 ± 0.95b 40 8 ± 0.75c 40 20 ± 0.55a <0.001
28-d BW loss, % 35 6 ± 1.20b 38 7 ± 0.94b 32 16 ± 1.11a <0.001
Means within rows with different letters are different (P < 0.05).
a–c

WB = whole-grain barley method; Zn = high dietary Zn method (containing 10,000 mg/kg of Zn as ZnO); CAL = California
1

method.

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All hens were provided the layer diets ad libitum
and received 16 h of light/d before the molting
programs began. The experimental period was
23 wk, consisting of a 1-wk preexperiment pe-
riod and 4 wk during the molt period, followed
by 18 wk of production.
The 3 molting programs were whole-grain
barley (WB), high dietary Zn (Zn), and Califor-
nia (CAL; Table 1). In the WB method, the hens
were fed whole-grain barley during the first 10
d and light was reduced to 8 h/d. On d 11, hens
were provided 100 g/d of the layer diet until d
28. On d 29, hens were provided the layer diet
ad libitum and the day length was increased to
16 h of light/d. In the Zn method, hens were fed
Zn diets containing 10,000 mg/kg of Zn as ZnO
for 10 d and light was reduced to 8 h/d. Hens
were fed 100 g/bird per day of the layer diet
from d 11 to 28. On d 29, hens were provided
the layer diet ad libitum and the day length was
increased to 16 h of light/d. In the CAL method,
hens were fasted during the first 10 d and light
was reduced to 8 h/d. On d 11, hens consumed
a cracked corn diet ad libitum until d 28. On d
29, hens were fed a layer diet ad libitum and
the day length was increased to 16 h of light/d.
The ingredients and chemical composition of
the diets are given in Table 2. Water was given
freely in all the groups during the experiment.
Traits Measured
All hens were weighed individually at the
beginning of the experiment and at 10 and 28
d following the start of the molt programs. One-
week preexperimental egg production, feed in-
take, and FCR, and 3-d preexperimental egg
weight were used to determine their initial val-
ues. Percentage of BW loss was calculated. Mor-
tality was recorded daily. Eggs were collected
daily then egg production was calculated on a
hen-day basis, and uncracked eggs were
weighed. Feed intake was measured weekly and
FCR (grams of feed:grams of egg) was calcu-
lated weekly during the postmolt period. Twenty

Table 4. Effects of different induced molting programs at different periods of the experiment on hen-day egg
production (%)

Molting program1

WB Zn CAL
Time of
experiment n Mean ± SEM n Mean ± SEM n Mean ± SEM P-value

Initial 40 47.5 ± 3.8 40 48.3 ± 2.9 40 50.0 ± 4.6 NS

Days 1–10 40 21.5 ± 1.8b 40 32.0 ± 2.8a 40 17.8 ± 1.5b <0.001
Days 11–28 35 17.6 ± 1.9b 38 27.5 ± 3.1a 32 0.7 ± 0.4c <0.001
Weeks 5–22 34 51.8 ± 2.6 33 43.0 ± 3.4 29 46.4 ± 2.4 NS

Means within rows with different letters are different (P < 0.05).
a–c

WB = whole-grain barley method; Zn = high dietary Zn method (containing 10,000 mg/kg of Zn as ZnO); CAL = California
1

method.
532 JAPR: Research Report

Table 5. Effects of different induced molting programs at different periods of the experiment on egg weight (g)

Molting program1

WB Zn CAL
Time of
experiment n Mean ± SEM n Mean ± SEM n Mean ± SEM P-value

Initial 47 60.96 ± 1.19 43 61.94 ± 0.86 62 62.87 ± 0.93 NS

Days 1–10 78 56.97 ± 0.96b 113 60.81 ± 0.61a 71 57.70 ± 0.75b <0.001
Days 11–28 92 60.74 ± 0.49 174 60.19 ± 0.94 0 — NS
Weeks 5–22 2,173 63.09 ± 0.14b 1,683 62.70 ± 0.13c 1,630 63.88 ± 0.14a <0.001

Means within rows with different letters are different (P < 0.05).
a–c

WB = whole-grain barley method; Zn = high dietary Zn method (containing 10,000 mg/kg of Zn as ZnO); CAL = California
1

method.

eggs per treatment were collected randomly to Antibody production against SRBC was

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determine the egg quality characteristics on the
first day of the 14th and 15th week of postmolt
production. The following characteristics were
recorded for the individual eggs: weight, shape
index, eggshell breaking strength, shell thick-
ness, yolk height and width, and albumen height,
width, and length. With these values, the yolk
index, albumen index, and Haugh units were
calculated. The egg quality analyses were com-
pleted within 24 h of collection.
On d 10 of the experiment, blood samples
were taken from the brachial vein of 17 hens in
each treatment and placed in tubes containing
EDTA to estimate the plasma corticosterone lev-
els. The bleeding procedure was limited to 1
min or less to minimize the influence of handling
stress [17]. All blood samples were collected at
the same time in the morning and centrifuged.
Plasma was frozen (−20°C) until analyzed for
the determination of corticosterone. Plasma cor-
ticosterone levels were measured by using kits
[18] and a gamma counter [19].
measured from 10 hens in each treatment at the
beginning of the experiment, and at 10 and 30
d following the start of the molting programs.
Hens were injected with 0.1 mL of 0.25% SRBC
in 0.9% saline. Circulating anti-SRBC antibody
titers were determined by the microhemaglutina-
tion technique from samples taken at 5 d after
the immunization. All titers were expressed as
the log2 of the reciprocal of the serum dilution
[20].
Nutrient analyses were determined ac-
cording to standard AOAC procedures [21]. The
analyses of Ca [22] and P [23] were done after
dry combustion of the diets. Metabolizable en-
ergy levels were estimated by using a prediction
equation [24]:
ME, kcal/kg = 53 + 38 [(CP, %) + (2.25
× EE, %) + (1.1 × starch, %) + (sugar, %)].
Statistical Analyses
Data were analyzed with SPSS for Windows.
Differences among the means of the molt treat-

Table 6. Effects of different induced molting programs on feed intake (g/hen per d) and feed conversion ratio (g
feed:g egg)

Molting program1

WB Zn CAL
Time of
experiment Item n Mean ± SEM n Mean ± SEM n Mean ± SEM P-value

Initial Feed intake 40 114.63 ± 0.77 40 115.10 ± 0.92 40 113.98 ± 0.98 NS

Feed efficiency 40 4.80 ± 0.28 40 4.36 ± 0.23 40 4.58 ± 0.26 NS
Weeks 5–22 Feed intake 34 111.68 ± 0.91b 33 110.24 ± 0.88b 29 115.74 ± 0.89a <0.001
Feed efficiency 34 3.79 ± 0.25b 33 5.24 ± 0.48a 29 4.46 ± 0.40ab 0.029

Means within rows with different letters are different (P < 0.05).
a,b

WB = whole-grain barley method; Zn = high dietary Zn method (containing 10,000 mg/kg of Zn as ZnO); CAL = California
1

method.
ONBAŞıLAR AND EROL: MOLTING METHODS AND PERFORMANCE 533

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Figure 1. Daily hen-day egg production (%) during the molt period. WB = whole-grain barley method; Zn = high
dietary Zn method (containing 10,000 mg/kg of Zn as ZnO); CAL = California method.

ment groups were studied by one-way ANOVA.
Means that were significantly different were sep-
arated by Duncan’s test. The frequencies of mor-
tality among groups were compared by using a
chi-square test [25].
RESULTS AND DISCUSSION
In this experiment, BW loss was signifi-
cantly higher in the CAL treatment than in the
other treatments (Table 3). Body weight loss is
an important factor contributing to the success
of induced molting because of its impact on
reproduction and on fat deposits [26, 27]. This
leads to regression of the reproductive tract,
which is proportional to the loss of BW, the
rejuvenation of the reproductive tract, and the
removal of fat accumulation, thereby increasing
tissue efficiency [28]. Webster [6] reported that
targeted weight losses have ranged from 15 to
35%, but the optimal BW loss of hens during
an induced molt was not identified.
In the present study during the molt period,
hen-day egg production was higher in the Zn
treatment than in the others (Table 4 and Figure
1). Postmolt (wk 5 to 22) hen-day egg production
was not significantly different among the 3 molt
treatments (Table 4 and Figure 2). This result
agrees with the findings of Park et al. [27], who
found that egg production was not different
among molting methods (feed withdrawal, Zn
acetate, Zn propionate). However, Alodan and
Mashaly [28] reported that egg production was
significantly affected by the molting program
[Zn, CAL, and on-again, off-again (ON-OFF)],
and the Zn group had significantly (P < 0.05)
lower egg production than the CAL and ON-
OFF treatments. In the current study, postmolt
egg production was lower than expected in all

Table 7. Effects of different induced molting programs at different periods of the experiment on antibody production
against sheep red blood cells1

Molting program2

Time of experiment WB Zn CAL P-value

Initial 2.6 ± 0.5 4.4 ± 0.8 4.6 ± 0.8 NS

Day 10 3.2 ± 0.6a 2.6 ± 0.2a 1.4 ± 0.2b 0.006
Day 30 5.7 ± 0.4a 5.9 ± 0.4a 4.6 ± 0.3b 0.043

Means within rows with different letters are significantly different (P < 0.05).
a,b
1
Values are log2 of reciprocal dilution of antibody titer.
2
WB = whole-grain barley method; Zn = high dietary Zn method (containing 10,000 mg/kg of Zn as ZnO); CAL = California
method. Values are means ± SEM (n = 10).
534
Figure 2. Weekly postmolt hen-day egg production (%) from wk 5 to 22. WB = whole-grain barley method; Zn =
high dietary Zn method (containing 10,000 mg/kg of Zn as ZnO); CAL = California method.
groups. These results showed that the genotype
(obtained by Ankara Poultry Research Institute)
used in this experiment was not suitable for the
second production cycle.
Postmolt egg weight was significantly higher
in the CAL group than in the other groups (Table
5). Contradictory to our findings, Alodan and
Mashaly [28] reported that different induced
molting programs (Zn, CAL, ON-OFF) had no
effect on egg weight. Previous studies of molting
methods have often shown conflicting results in
hen responses because of different conditions
[4]. Body weight at the start of the experiment,
BW loss [29], age of the hens [26], and strain
[30] have all been shown to affect the results
of different molting methods. There were no
significant differences among treatments in the
egg quality characteristics examined in the cur-
rent study (data not shown). Similarly, Alodan
and Mashaly [28] showed that Zn methods did
not affect egg quality. McCormick and Cunning-
ham [10] also reported that egg quality variables
and shell breaking strength were not signifi-
cantly different between different molting meth-
ods (fasting for 10 d and high dietary Zn).
Hens in the WB and Zn treatments consumed
much less feed (P < 0.001) from the 5- to 22-
wk postmolt than hens in the CAL group (Table
JAPR: Research Report
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6). The FCR of hens in the WB treatment was
better than that of hens in the Zn group. Berry
and Brake [7] reported that hens in the low di-
etary Na treatment had significantly better (P <
0.05) feed efficiency than those in the high di-
etary Zn and fasting treatments.
During this experiment, 6 (15%), 7 (17.5%),
and 11 (27.5%) hens died in the WB, Zn, and
CAL treatment, respectively. Mortality was not
significantly affected by the different molting
treatments. This result is in agreement with Alo-
dan and Mashaly [28].
One method of assessing welfare in different
molting programs is to measure the physiologi-
cal parameters associated with the stress re-
sponse. Physiological parameters of stress in
poultry, as reviewed by Freeman [31], Siegel
[32], and Hill [33], may include increases in
circulating levels of corticosterone and suppres-
sion of humoral immunity. In this experiment,
plasma corticosterone levels (mean ± SEM) of
hens in the WB, Zn, and CAL treatments were
determined as 3.78 ± 0.16, 4.07 ± 0.13, and 4.71
± 0.19 ng/mL, respectively. The higher (P =
0.001) corticosterone levels in the CAL treat-
ment could reflect a period of higher-stress con-
ditions than in the Zn and WB treatments. How-
ever, Keshavarz and Quimby [34] reported that
ONBAŞıLAR AND EROL: MOLTING METHODS AND PERFORMANCE 535

corticosterone levels were not significantly dif-
ferent among various molting regimens (feed
withdrawal, continuous corn diet, and feed with-
drawal-corn diet groups).
On d 10 and 30 of the experiment, antibody
titers to SRBC in the CAL method were signifi-
cantly lower than in the other treatments (Table
7). This reduction may be due to the increase in
circulating levels of corticosterone. Corticoste-
rone can reduce the antibody response to an
antigen in birds [35]. Nathan et al. [36] showed
that short-term starvation diminished the re-
sponse to SRBC. Conversely, Alodan and Mas-
haly [28] showed that the different induced molt-
ing programs did not adversely affect antibody
production against SRBC. Some of these differ-
ences may be due to the genetic background of
the birds, the different molting programs, or the
use of different suspensions of sheep eryth-
rocytes.

CONCLUSIONS AND APPLICATIONS

1. In the CAL method, egg weights and plasma corticosterone levels were higher and humoral

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immunity was lower than with other methods. Hens molted with this method also had greater
feed consumption than with the other methods; however, egg production was not affected.
Because of physiological stress, this molting program may be less suitable for the welfare of
laying hens.
2. When the WB and Zn methods were compared, egg weight with the WB method was higher
and FCR was better than with the Zn method. Egg production, mortality, egg quality, plasma
corticosterone levels, and immunity of the laying hens were not significantly different between
these 2 treatments.
3. Therefore, in this study the WB molting program resulted in the best production performance
among the programs examined.

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