Effects of a phytogenic feed additive on growth performance, selected blood criteria and jejunal morphology in broiler chickens

Abstract

The study was conducted to examine the effects of a phytogenic feed additive (PFA; BIOSTRONG® 510) on growth performance, selected blood parameters and jejunal morphology in broiler chickens. The PFA consists of a mixture of essential oils with thymol and anethole as leading active substances, as well as different herbs and spices. A total of 264 1-d-old Cobb male broilers were randomly allocated to 2 dietary treatments, with 6 replicates per treatment and 22 birds each replicate. The experiment lasted 42 days. The dietary treatments were a starter and grower diet without feed additives (control), or the diets supplemented with 150 mg/kg of the PFA. Body weight and feed intake were not significantly influenced by PFA feeding compared with the control during all experimental periods. During the grower phase (22 - 42 d of age) and during the whole period (1 - 42 d of age) the PFA significantly improved (P<0.05) feed conversion ratio. Serum total protein, albumin, total cholesterol and leucocytes were increased by PFA feeding. Furthermore, villus height to crypt depth ratio in the jejunum was increased by PFA feeding. In conclusion, the results of this study show that inclusion of PFA lead to morphological changes in the jejunum, which might influence nutrient absorption and thus, improved FCR.

Full text

Emir. J. Food Agric. 2013. 25 (7): 549-554
doi: 10.9755/ejfa.v25i7.12364
http://www.ejfa.info/
549
ANIMAL SCIENCE
Effects of a phytogenic feed additive on growth performance, selected blood
criteria and jejunal morphology in broiler chickens
Abdulkarim A. Amad1*, K. R. Wendler2and J. Zentek3
1Department of Animal Production, Faculty of Agriculture and Veterinary, Thamar University,
Dhamar, Yemen
2Delacon Biotechnik, Steyregg, Austria
3Institute of Animal Nutrition, Department of Veterinary Medicine, Freie Universität Berlin, Berlin, Germany
Abstract
The study was conducted to examine the effects of a phytogenic feed additive (PFA; BIOSTRONG® 510) on
growth performance, selected blood parameters and jejunal morphology in broiler chickens. The PFA consists
of a mixture of essential oils with thymol and anethole as leading active substances, as well as different herbs
and spices. A total of 264 1-d-old Cobb male broilers were randomly allocated to 2 dietary treatments, with 6
replicates per treatment and 22 birds each replicate. The experiment lasted 42 days. The dietary treatments were
a starter and grower diet without feed additives (control), or the diets supplemented with 150 mg/kg of the PFA.
Body weight and feed intake were not significantly influenced by PFA feeding compared with the control
during all experimental periods. During the grower phase (22 - 42 d of age) and during the whole period (1 - 42
d of age) the PFA significantly improved (P<0.05) feed conversion ratio. Serum total protein, albumin, total
cholesterol and leucocytes were increased by PFA feeding. Furthermore, villus height to crypt depth ratio in the
jejunum was increased by PFA feeding. In conclusion, the results of this study show that inclusion of PFA lead
to morphological changes in the jejunum, which might influence nutrient absorption and thus, improved FCR.
Key words: Broiler chicken, Performance, Blood parameters, Jejunal morphology
Introduction
The ban of antibiotic growth promoters (AGP)
in poultry feeds intensified the search for
alternatives improving the health and productivity
of broiler chickens (Barreto et al., 2008). Such
alternatives are probiotics, prebiotics and organic
acids, which are added to the feed (Huyghebaert et
al., 2011). Also phytogenic feed additives (PFA)
were shown to enhance performance in AGP-free
livestock production (Alçiçek et al., 2003; Steiner,
2009). Phytogenic feed additives consist of a broad
variety of substances, mainly extracts from plant
materials, such as flowers, buds, seeds, leaves,
twigs, bark, herbs, wood, fruits and roots (Burt,
2004). The active molecules include many different
secondary plant metabolites, resulting in a broad
range of physiological effects, like secretolytic and
spasmolytic, or immune-stimulative effects (Lee et
al., 2004a).
PFA are generally recognized as natural feed
additives and safe to the animal. However, results
of studies concerning the use of PFA in broiler
nutrition are inconsistent (Windisch et al., 2008).
Some authors stated significant improvement of
broiler performance (Ertas et al., 2005; Cross et al.,
2007), whereas others reported no effects on BW
gain and feed intake (Lee et al., 2003; Jamroz et al.,
2005; Nasir and Grashorn, 2010) or feed
conversion ratio (Ocak et al., 2008). These
discrepancies may be due to numerous factors such
as type and parts of plants used, their physical
properties, time of harvest, the preparation method
of PFA and their compatibility with other feed
components (Jang et al., 2007). Furthermore, the
mode of action of these additives is not fully
clarified yet and in vivo studies are limited. Plant
extracts have been shown to influence digestion and
secretion of digestive enzymes (Platel and
Srinivasan, 2000; Williams and Losa, 2001) to
increase absorption of micronutrients (Usha et al.,
2010) and to exhibit antibacterial, antiviral and
Received 14 May 2012; Revised 29 December 2012
; Accepted
08 January 2013; Published Online 01 May 2013
*Corresponding Author
Abdulkarim A. Amad
Department of Animal Production, Faculty of Agriculture and
Veterinary, Thamar University, Dhamar, Yemen
Email: al_absie@yahoo.com

Abdulkarim A. Amad et al.
550
antioxidant activities (Brenes and Roura, 2010).
However, only few studies investigated the effects
of PFA on intestinal morphology in broiler
chickens. Therefore, the aim of the present study
was to examine the effect of a commercial PFA on
performance, intestinal morphology and selected
blood parameters in broiler chickens.
Materials and Methods
Two hundred sixty four 1-d-old male Cobb
chickens were weighed and randomly allotted to 2
experimental treatments. Each treatment consisted
of 6 replicates with 22 birds per replicate. The
dietary treatments were a starter and grower basal
diet without any feed additives used as a control or
the basal diets supplemented with 150 mg/kg of a
commercial PFA (BIOSTRONG®510, Delacon,
Steyregg, Austria) used as experimental diets. The
PFA consisted of a mixture of essential oils, with
thymol and anethole as leading active substances,
as well as different herbs and spices. The
compositions of the basal diets are shown in Table
1. Control and experimental diets were formulated
to be iso-nutritive and to meet the requirements for
broiler chickens during starter and grower phase.
Feed and water were available ad libitum. Feed
consumption and BW of the birds were recorded
weekly and were used to calculate broiler
performance (weight gain, feed intake, feed
conversion ratio).
Blood samples were taken at 35 d of age from
the brachial vein from 12 birds per treatment group
(2 birds per pen). Blood samples were separated by
centrifuge, and serum was analyzed for glucose,
total cholesterol, triglycerides, albumin and total
protein using commercial test kits. Blood cell
counts were determined by commercial cell
counter.
At 42 d of age, 9 birds per treatment were
slaughtered and the jejunum was removed for
assessment of tissue morphology. The tissue
samples were taken from the middle part of the
jejunum (from the entrance of bile duct to Meckel’s
diverticulum, pieces about 5 cm in length),
carefully cleansed and then fixed in 4% buffered
formalin for 2 days. The further processing
consisted of serial dehydration in PBS and graded
ethanol solutions, clearing with xylene and
embedding in paraffin. Sections of 5 µm were
prepared and placed on glass slides. Tissue samples
were deparaffinised, rehydrated and stained with
haematoxylin and eosin. Villus heights and crypt
depths were examined with a photomicroscope
(Photomikroskop III, Carl Zeiss, Oberkochern,
Germany) fitted with a digital camera (MikroCam 3
MP, Bresser, Rhede, Germany) and images were
analysed using image analysis software (Image
software Bresser MikroCamLab Mikroskopie). A
total of 16 intact well-oriented villus-crypt units
were randomly selected at each tissue sample.
Villus height was measured from the tip of the
villus to the villus-crypt junction, and crypt depth
was defined as the depth of the invagination
between two villi.
Experimental data were analyzed by ANOVA
using SPSS v. 17.0 (Statistical Packages for the
Social Sciences, released August 23, 2008). Data
was tested for homogeneity of variances, and
comparison of means was performed by Tukey test.
The significance level was set at p <0.05.
Results
During the whole experiment, birds were
healthy and the mortality was below 1%.
Performance data for broiler chickens during the
starter and grower phase and for the total
experimental period are summarized in Table 2.
Table 1. Ingredients and nutrient composition of
experimental diets.
Ingredients (%) Starter
1 - 21d
Finisher
22 - 42 d
Soybean meal
(
48%
CP
)
34.50
31.80
Maize 28.46 29.47
Wheat 24.69 24.29
Soy oil 6.80 9.35
Limestone
1.80
1.51
Monocalcium-phosphate 1.40 1.35
Premix* 1.20 1.20
Chromium oxide 0.50 0.50
DL
-
Methionine
0.30
0.29
L-Lysine 0.25 0.14
Additives** 0.10 0.15
100% 100%
Chemical composition (calculated)
AMEn, (MJ/k
g)
12.59
13.29
Crude protein( %) 22.89 21.50
Crude fibre (%) 2.39 2.32
Lysine (%)
1.43
1.26
Methionine (%)
0.64
0.61
Methionine + Cystein (%) 1.02 0.98
Calcium (%) 1.03 0.90
Total Phosphorus (%)
0.70
0.68
*Supplied per kg of diet: 4000 IU vitamin A (retinyl acetate); 400 IU cholecalciferol; 80
mg (α-tocopherole acetate); 3 mg vitamin K3 (menadione); 2.5 mg thiamin ; 2.5 mg
riboflavin; 25 mg nicotinic acid; 4 mg pyridoxine; 0.02 mg cobalamin; 0.3 mg biotin; 10
mg calcium pantothenate acid; 1 mg folic acid; 800 mg choline chloride; 50 mg Zn (Zinc
oxide); 20 mg Fe (Iron carbonate); 60 mg Mn (manganese oxide); 12 mg Cu (copper
sulfate-pentahydrate); 0.45 mg J (calcium iodate; 0.30 mg Co (cobalt- (II)-sulfate-
heptahydrate); 0.35 mg Se (sodium selenite); 1.3 g Na (sodium chloride); 0.55 g Mg
(magnesium oxide) ; ** Additives: Control = without active substances (PFA); Treatment
= Biostrong® 150 mg/kg feed.

Emir. J. Food Agric. 2013. 25 (7): 549-554
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551
For all experimental periods, there was no
effect (P>0.05) of the PFA on BWG and feed
intake. Feed conversion ratio was significantly
improved (P<0.05) in the grower phase (22 - 42 d
of age) as well as during the whole experimental
period (1- 42 d of age) due to PFA
supplementation.
Villus heights, crypt depths and villus height:
crypt depth ratios of jejunal tissue samples are
presented in Table 4. In broilers fed diets
supplemented with PFA, there were no differences
between the villus height and crypt depth could be
observed compared to the control (Figure 1). Villus
height: crypt depth ratio was significantly (P<0.05)
increased in birds fed PFA compared to the control.
Effect of the phytogenic feed additive on some
blood parameters in broilers were summarized in
Table 3. Serum total protein, albumin, cholesterol
and leucocytes were increased (P<0.05), while
serum glucose, triglycerides, hemoglobin and
erythrocytes were not affected by PFA feeding.
Table 2. Effect of the phytogenic feed additive on broiler performance during the starter and grower period (means ± SD).
ab Values with different superscript within lines differ significantly (P<0.05)
Table 3. Effect of the phytogenic feed additive on some blood parameters in broilers (35th d of age; means ± SD).
Parameters Treatments P-value
Control PFA (150 mg/kg)
Serum total protein (g/l) 27.03
a
±3.65 30.64
b
±1.38 <0.004
Serum albumin (g/l)
11.67
a
±2.15
14.22
b
±0.87
0.001
Serum glu
cose (mmol/l)
14.22
±1.49
14.99
±0.60
0.11
Serum Cholesterol (mmol/l) 3.00
a
±0.65 3.84
b
±0.22 <0.001
Triglycerides (mmol/l) 1.33 ±1.23 1.86 ±0.71 0.21
Erythrocytes
(Tpt/l)
2.27
±0.10
2.34
±0.27
0.31
Leucocytes
(Tpt/l)
175.8
a
±2.4
183.5
b
±9.5
0.012
Haemoglobin (mmol/l) 4.94 ±0.25 5.22 ±0.48 0.09
ab Values with different superscript within lines differ significantly (P<0.05)
Table 4. Effect of the phytogenic feed additive on histomorphological parameters of the jejunum in broilers (means ± SD).
Parameters
Treatments
P- value
Control
PFA
(
150
mg
/
kg
)
Villus height (µm)
1447±125.4
1569±120.9
0.068
Crypt depth (µm)
199±21.0
179±18.6
0.061
Villus height:crypt depth 7.3
a
±0.8 8.8
b
±0.9 0.004
ab Values with different superscript within lines differ significantly (P<0.05)
Period
Treatments
P-value
C
ontrol
PFA
(
150
mg
/
kg
)
Average body weight
gain (g)
1
-
21
d
797
±67
813
±28
0.62
22 - 42 d 2108 ±102 2144
±150 0.64
1 - 42 d 2906 ±134 2956 ±173 0.58
Average feed intake (g)
1 - 21 d 1124 ±108 1106 ±99 0.76
22
-
42
d
3332
±130
3259
±211
0.49
1
-
42
d
4456
±202
4365
±287
0.54
Feed
conver
sion ratio (g/g)
1
-
21
d
1.41
±0.07
1.36
±0.08
0.295
22 - 42 d 1.58
a
±0.04 1.52
b
±0.03 0.013
1 - 42 d 1.53
a
±0.04 1.48
b
±0.02 0.011

Abdulkarim A. Amad et al.
552
Figure 1. Villus height and crypt depth in the jejunum of broilers fed diets with or without the phytogenic feed additive at
42 days of age, left control, right phytogenic feed additive.
Discussion
In the present study, there was no effect of the
tested PFA on BW and feed intake in broilers.
These results are consistent with the studies of Jang
et al. (2007) and Erdogan et al. (2010), who did not
find effects of different phytogenic compounds on
broiler growth performance. However, FCR was
significantly improved in the present study.
Consistently, Jamroz et al. (2005) reported
improved FCR due to the addition of a plant
extract, containing cinnamaldehyde, carvacrol and
capsaicin, to a maize or wheat and barley based diet
by 4.1 or 2.0%, respectively, whereas BW was not
affected by treatment. Al-Kassie (2009) showed
that the addition of 200 ppm oil extract derived
from thyme and cinnamon to broiler diets
significantly improved BW gain and FCR during a
growing period of 6 weeks and Ocak et al. (2008)
reported higher (P< 0.05) BW at 21 and 42 d of age
as well as higher (P< 0.05) BW gain from 7 to 35
days of age in broilers fed peppermint and thyme
compared to the controls. The above mentioned
studies, reporting improved FCR, have in common,
that the added substances mainly consist of
essential oils. Essential oils exert antimicrobial
activity in the digestive tract of animals (Lee et al.,
2004; Al-Kassie, 2010). It is hypothesized that gut
microflora reduces nutrients available to the animal
by enforcing the intestinal cell turnover and thereby
increasing the intestinal requirement for nutrients to
maintain tissue integrity. Moreover, intestinal
microflora and epithelial cells have to compete for
nutrients (Dibner and Richards, 2005; Lan et al.,
2005). By a reduction of intestinal microflora,
essential oils may lead to moderate cell turnover, to
decreased intestinal nutrient requirements and to
less competition for available nutrients. As a
consequence, FCR is improved because more
nutrients are used for BW gain instead of tissue
maintenance or microbial growth.
The increased nutrient supply for growth is
reflected in enhanced nutrient transport in the
blood. For example, Ghazalah and Ali (2008)
observed higher levels of total protein, albumin and
globulin in the blood serum of birds when fed 0.5%
rosemary leaves. Similar results were obtained in
the present study, where higher contents of total
protein and albumin in the blood serum of PFA fed
animals may also indicate enhanced nutrient supply
and transport. Furthermore, Tekeli et al. (2006)
reported increased blood glucose concentration by
Zingiber officinale supplementation and increased
triglyceride concentration by both Zingiber
officinale and Syzygium Aromaticum
supplementation. In the present study, there was no
effect of PFA on blood glucose and triglyceride
concentrations. Calislar et al. (2009) found effects
neither on triglycerides nor on blood cholesterol
levels by application of a PFA containing
Origanum vulgare ssp. hirtum extract. In general,
essential oils are more often associated with
hypocholesterolemic properties (Lee et al., 2004).
In contrast, in the present study chicken fed the
PFA showed higher serum cholesterol
concentrations compared to the control animals.
This discrepancy may be due to the combination of
essential oils and pungent substances which were
also included in the used PFA. Pungent substances
increase digestive secretions including enzymes and
bile. The higher serum cholesterol content, which
was observed in the PFA fed animals of the present
study, may be the result of an increased lipid
digestibility due to a higher secretion of bile and
digestive enzymes. The improvement of FCR as

Emir. J. Food Agric. 2013. 25 (7): 549-554
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well as the increase of blood nutrient concentrations
supports this assumption, although it remains
speculative as nutrient digestibilities have not been
investigated in the present study.
Regarding the intestinal morphology, heavier
chickens are generally associated with longer villi,
greater villus width and higher villus surface area
as compared to lighter ones (Adibmoradi et al.,
2006; Incharoen et al., 2010). In the present study
there were insignificantly increased villus heights
and decreased crypt depths in the jejunum of birds
receiving 150 mg/kg PFA compared to the control.
Accordingly, the villus height: crypt depth ratio
was significantly higher in the birds fed the PFA
compared to the control. These findings were
consistent to Adibmoradi et al. (2006) who reported
that jejunal villus height was increased whereas
crypt depths were decreased, leading to increased
villus height: crypt depth ratio in birds fed graded
levels of garlic meal. It has been suggested that
longer villi would result in an increased surface
area and higher absorption of available nutrients
(Caspary, 1992; Yasar and Forbes, 1999). A higher
absorptive capacity of the intestine of PFA fed
animals is also supported by higher blood nutrient
concentrations of those animals, as observed in the
present study. However, although Jamroz et al.
(2006) reported similar improvement of FCR due to
the supplementation with a plant extract containing
carvacrol, cinnamaldehyde and capsicum oleoresin,
the authors could not find an effect of the plant
extract on intestinal morphology in broilers at 42 d
of age. Thus, for PFA containing essential oils as
well as pungent substances further, eventually
synergistic, effects or other modes of action cannot
be ruled out.
Conclusion
In the present study, the addition of the
phytogenic feed additive BIOSTRONG® 510 to
broiler diets significantly improved feed conversion
ratio. The observed improvement of feed
conversion ratio may be caused by a combination
of different effects, including enhancement of
digestive secretions, antimicrobial effects of
essential oils as well as enlargement of intestinal
absorptive surface. However, further studies are
necessary to evaluate the effects of various
substances present in phytogenic feed additives and
to clarify their specific modes of action.
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