Efficacy of Allicin Supplementation on Zootechnical Performance and Immunological
Parameters of Broiler Chickens
M. A. Tony1*, Samah H. Mohamed2, Ashgan F. El-Sissi2 and Abeer H. Abdel Razek3
1) Department of Nutrition and Clinical Nutrition, Faculty of Veterinary Medicine, Cairo
University, Egypt (*Corresponding author E-mail: [email protected])
2) Department of Immunology, Animal Health Research Institute, Dokki, Giza, Egypt
3) Department of Veterinary Hygiene and Management, Faculty of Veterinary Medicine,
Cairo University, Egypt
Abstract
An experiment was conducted to assess the effect of feed supplementation of allicin powder
on growth performance and immune response of broiler chickens. One hundred eighty 1-dold Hubbard chicks of both sexes were randomly assigned to 3 floor pens (60 chicks/pen).
Group A consumed basal diets (starter and grower-finisher) and served as a control group.
Group B and C were fed on the same basal diets supplemented with 125 and 250 g Allicin
25%® per ton feed respectively. Feed and water were provided ad-libitum during 35 days
experimental period. Body weights as well as the rest of feed were recorded weekly. Body
weight gain and feed conversion were calculated. Blood samples were collected according to
a time program to evaluate some immunological parameters include antibodies titers against
vaccines used. At the end of the experiment, ten birds were randomly selected from each
group to evaluate carcass yield. The results reported that the two levels of allicin used had
significant effects (p<0.05) on feed intake, weight gain and feed conversion from the third
week of age till the end of the experimental period compared with the control group. The
results of immunological parameters measured revealed that allicin treatments could enhance
broiler chickens innate immunity as it significantly increased (p<0.05) phagocytic activity
and humoral immune responses against vaccines used. In addition, allicin treatment
maintained oxidant/antioxidant balance. Carcass weights, dressing percent, carcass cuts and
some organs weights were not affected by allicin supplementation. However, abdominal fat
content was reduced significantly (P<0.05) in both supplemented groups. In conclusion,
using allicin as a feed additive could improve broiler chicken performance, reduce abdominal
fat deposition and enhance immune status.
Key words: Broiler chickens, Allicin, Performance, Immune responses.
Introduction
Allicin is an organosulfur compound obtained from garlic, a species in the family Alliaceae
(Eric B., 1985). Allicin has a distinctively pungent smell and exhibits antibacterial, antifungal, anti-inflammatory and antioxidant properties (Sela et al., 2004; Bautista et al., 2005
and Lindsey et al., 2005).The use of antibiotics as growth promoters is facing serious
criticism. There are some important reasons that restrict the use of antibiotics such as the drug
resistance in bacteria and the drug residues in meat. To overcome the poor performance and
the increase susceptibility to diseases resulted from removal of antibiotics from birds diets,
attempts were made to find other alternatives. The utilization of growth promoters of natural
origin became of an interest in recent years (Iji et al., 2001). Vaidya et al. 2009 and Block
et al. 2010 clarified the mechanism of the antioxidant or anti-stress activity of allicin, such as
trapping free radicals. When allicin decomposes, it forms 2-propene sulfenic acid, and this
compound is what binds to the free-radicals as an anti-stress agent. Allicin has been found to
lower serum and liver cholesterol (Qureshi et al.,1983), inhibit bacterial growth (Cavallito et
al., 1994) and reduce oxidative stress (Lindsey et al., 2005 and Choudhary, 2008). Also
allicin has immune-stimulatory effect (Cho et al 2006).
In broilers, it was reported that garlic, as a natural feed additive, improved broiler growth and
feed conversion ratio (FCR), and decreased mortality rate (Tollba et al., 2003). Amagase et
al., (2001) and Demir et al., (2003) reported that improvement of broilers performance and
carcass merits can be achieved by supplementation of diets with garlic powder. Demir et al.,
(2003) recognized that the strong stimulating effect of garlic on the immune system of
broilers is mainly attributed to the bioactive components of garlic including sulphur
containing compounds such as allin, and Allicin. However, other studies have shown a
strong oxidative effect in the gut that can damage intestinal cells in laboratory animals.
Furthermore, many of these results were obtained by excessive amounts of allicin, which has
been clearly shown to have some toxicity at high amounts, or by physically injecting the
lumen itself with allicin, which is not indicative of what would happen via oral ingestion of
allicin or garlic supplements (Banerjee et al., 2001).
Allicin effects on broilers performance, carcass characteristics and immune status are
debatable. The objective of this study was to investigate the effect of feeding allicin by two
different doses on performance and immune status of broiler chickens.
Materials and Methods
Allicin 25%®: is a commercial powder feed additive product.Allicin 25%®manufactured by
Anhui Ruisen Bio-Tech Co., LTD, China. The recommendation for use as feed additive for
poultry is 125 g per ton feed.
Experimental Birds, houses and diets
A total number of 180 day-old chicks Hubbard chicks of both sexes and average body weight
of 45 g ±2 g were randomly assigned into 3 floor pens (60 chicks/pen) in the experimental
house of Department of Veterinary Hygiene and Management, Faculty of Veterinary
Medicine, Cairo University, Egypt. Group A consumed the basal dies (starter and growerfinisher) and served as a control group. Group B was fed on the same basal dies
supplemented with 125 g Allicin 25%®per ton feed. Group C was reared on basal diets
supplemented with high dose of Allicin 25%®(250 g per ton feed).
Corn-soybean meal based diets were formulated to cover the nutrient requirements of
Hubbard broilers (Hubbard manual catalogue). Two stages diets (starter and grower-finisher)
in the form of mash and water were provided ad-libitum during the 35 days experimental
period (Table 1). The individual body weight for all birds as well as the rest of feed was
recorded weekly. Body weight gain and feed conversion were calculated.
Birds in all experimental groups were vaccinated against Newcastle and Gumboro diseases
according to the vaccination program showed in table (2).
Immunological parameters
Assay of phagocytosis:
Heparinised blood samples were collected (8 samples /group) at 5th day post 1st &2nd ND
(Bivalent Hitchner&Lasota) and Gumboro vaccination. They were used for determination of
phagocytic activity of peripheral blood using candida albicans according to Bos and Souza
(2000). The phagocytic activity was determined by phagocytic % (number of phagocytic
macrophages / total number of macrophages) and phagocytic index (number of macrophages
engulf ≥3 Candida spores /total number of phagocytic macrophages).
Table 1. Composition % and calculated nutrients profile of the basal diets
Ingredients %
Starter
Grower-finisher
(1 – 21 day)
(22 – 35 day)
yellow Corn
53.70
56.65
Corn gluten meal
5.19
4.6
Soybean meal (44% CP)
35.30
31.0
Soy oil
2.10
4.00
Bone meal
2.30
2.40
Limestone
0.50
0.45
Common salt
0.30
0.30
Sodium bicarbonate
0.10
0.10
DL-Methionine
0.16
0.15
L-Lysin
0.05
0.05
Vitamin & mineral premix*
0.30
0.30
Calculated analysis:
ME (Kcal/kg)
2950.0
3100.0
Crude Protein%
22.5
20.5
Crude fat%
5.8
6.92
Crude fiber%
5.0
4.8
Calcium%
1
1
Non-phytate phosphorus%
0.45
0.45
Chloride%
0.22
0.22
Sodium%
0.18
0.18
Methionine%
0.51
0.51
Lysine%
1.14
1.03
Methionine+cystine%
0.90
0.90
* Per kg premix: 1 200 000 IU vit. A, 350 000 IU vit.D3, 4 000 mg vit.E, 250 mg
vit.B1, 800 mg vit.B2, 600 mg vit.B6, 3.2 mg vit.B12, 450 mg vit. K3, 4.5 g nicotinic
acid, 1.5 g Ca-pantothenate, 120 mg folic acid, 5 mg biotin, 55 g choline chloride, 3
g Fe, 2 g Cu, 10 g Mn, 8 g Zn, 120 mg I, 40 mg Co.
Table 2. Vaccination programme
Age (days)
Vaccine*
7
Bivalent Hitchner B1 + IB
14
Hitchner B1
16
Gumboro D78
21
Lasota
28
Gumboro D78
*Vaccines were obtained from Intervet, Inc., Egypt
Application
Eye dropping
Eye dropping
Drinking water
Drinking water
Drinking water
Lysozyme & Nitric oxide assays:
Serum samples collected at 1st day post 1st & 2nd Newcastle and Gumboro vaccines and at the
end of the experiment. Lysozyme activity was estimated by agarose gel plate lyses assay
according to Peeters and Vantrappen (1977) using Micrococcus lysodeikticus bacteria. The
concentration of lysozyme was obtained from logarithmic curve prepared using standard
lysozyme solution. Nitric oxide was carried out according to Yang et al., (2010) using Griess
reagent after removal of protein by mixture of ZnSO4 & NaOH. Conversion of nitrate to
nitrite using Cu plated Cd. NO concentration was calculated from standard curve using
NaNO2 Detection of Antibodies titers to NDV and IBDV vaccines. Serum samples were
collected weekly from all groups (15 samples/group), for detection of antibodies titers
against NDV using Haemagglutination inhibition test (HI) according to Beard, 1989 and
against Gumboro disease (IBDV) using ELISA kit according to the manufacture`s instruction
(GmbH, Germany). Malondialdehyde was measured chemically according to Ohkawa, (1979)
and glutathione was measured according to Ellman et al., (1959) at the end of the
experimental period.
Carcass characteristics
At the end of the experiment, ten birds /group were randomly selected from all groups. Feed
was withdrawn 12 h before slaughter. Birds were defeathered and eviscerated after
slaughtering by bleeding the jugular vein. The gizzard, heart, liver, small intestine, cecum and
abdominal fat were excised and weighed and their relative weights to live BW (%) were
calculated. After removal of head, shanks and offal, ready to cook carcass was obtained. The
ready to cook carcass weight was then determined, and the carcass yield percentage
(dressing%) was calculated. Weights of breast, thigh, back, wing and neck were recorded.
Statistical analysis
All data were statistically analyzed using IBM SPSS® version 19 software for personal
computer-2010. Means were compared by one way ANOVA (p<0.05) using Post Hoc test
and least significant difference (LSD) according to Petrie and Watson (1999).
Results and Discussion
Zootechnical performance
Production performance parameters of the three experimental groups are shown in table 3.
The two levels of allicin used had positive effects on feed intake, weight gain and feed
conversion from the third week of age till the end of the experimental period. These results
are in agreement with those of Javandel et al., 2008 and Lewis et al., (2003) who observed a
positive effect of garlic on performance of broiler chickens. Demir et al., (2003) reported an
improvement in body weight gain and feed conversion in broiler chickens fed low
concentrations of commercial garlic products. In contrast with these results, Horton et al.,
(1999) and Onibi et al., (2009), reported that supplementation of garlic powder had no
significant effect on weight gain and feed conversion. However increasing the dose of allicin
had no beneficial effect (p<0.05) on weight gain and feed intake. The best feed conversion
was recorded in group B which was fed on diets containing low dose of allicin. Thus the
results of this trial suggest the use of this form of allicin 25%® in a concentration of 125 g/ton
feed. The recorded results are in agreement with those recommended by Ari et al., 2012 who
reported that supplementation of garlic will lead to optimal reduction in cholesterol
deposition and supports performance of broilers through its antibacterial and antioxidant
effects.
Immunological parameters
The results of phagocytic activity (Table 4) showed a significant (p<0.05) increase in
phagocytic index and percent in groups B and C compared with control group at 5 days post
1st and 2nd vaccinations with IBDV and at 5 days post 2nd vaccination with NDV. The data
from this study are in agreement with Salman et al., (1999) and Kanget et al. (2001) who
found that allicin has stimulatory effect on phagpcytic activity of macrophages in vitro
studies. Also Sharma et al. (2010) and Talpur and Ikhwanuddin (2012) revealed that garlic
can increase the non specific immunity by activating phagocytosis in mice and fish
respectivity. The immunostimulatory activity of allicin may be mediated through
upregulation of secretory molecules in macrophages and also it plays a role in triggering the
activation of macrophages Kanget et al., (2001). Moreover allicin induced activation of
extracellular signal-regulated kinases 1and 2(ERK1/2) which belong to the mitogen-activated
protein kinase family leading to activation of macrophages Patya et al., (2004).
Table 3. Effect of allicin powder on performance parameters of broiler chickens
(means±SE)
Parameter
Period
Group A
Group B
Group C
(Control)
(Allicin125 gm) Allicin 250 gm )
Body weight
7d
145.16±0.21a
146.91±0.20a
146.89±0.22a
(g/bird)
14 d
471.18±0.27a
472.91±0.15a
473.08±0.18a
a
a
850.32±0.21
850.18±0.21
850.19±0.19a
21 d
a
b
28 d
1390.11±0.15
1408.91±0.25
1409.21±0.28b
1980.37±0.48a
2011.17±0.68b
2010.95±0.63b
35 d
a
a
Weight gain
7d
100.16±0.23
101.91±0.22
101.89±0.24a
(g/bird)
14 d
326.02±0.18a
326.00±0.20a
326.19±0.23a
a
b
379.14±0.33
377.27±0.24
377.11±0.26b
21 d
28 d
539.79±0.38a
558.73±0.23b
559.02±0.18b
a
b
35 d
590.26±0.45
602.26±0.31
601.74±0.22b
Total
1935.37±0.47a
1966.17±0.66b
1965.95±0.65b
a
a
Feed intake
7d
127.35±0.31
128.09±0.36
128.11±0.32a
(g/bird)
14 d
489.98±0.55a
495.63±0.56a
496.03±0.50a
a
a
546.95±0.38
550.15±0.44
560.03±0.61b
21 d
747.91±0.13a
751.88±0.33b
750.75±0.32b
28 d
a
b
35 d
1120.6±0.60
1163.5±0.68
1165.2±0.55b
Total
3032.79±0.75a
3089.25±0.65b
3100.12±0.45c
Feed conversion
7d
1.27
1.26
1.26
(FCR)
14 d
1.50
1.52
1.52
21 d
1.55
1.46
1.49
28 d
1.39
1.35
1.34
35 d
1.90
1.93
1.94
Total
1.57
1.57
1.58
Values in the same row with different superscripts are statistically significantly different
(p<0.05).
Table 4. Effect of the dietary supplementation of alicin on phagocytic % and index of
Peripheral blood mononuclear cells of broiler chickens.
Phagocytic percent
phagocytic indix
PARAMETERS
Group B
Group C
Group B
Group C
Group A
(Allicin 125 (Allicin 250
Group A
(Allicin 125 (Allicin 250
TIME
(control)
gm)
gm)
(control)
gm)
gm)
5th day post 1st NDV
60±2.91
59±4.62
57±4.91
5thday post 1st IBDV
64±1.20a
75±2.73b
77±2.58b
71±1.53
72±2.40b
0.58±0.027a
0.76±0.012b 0.76±0.009b
74±3.02
0.60±0.027a
0.11
0.69±0.015
5thday post 2nd NDV 62±2.34a
5thday post 2nd
IBDV
65±2.60
LSD
9.42
70±1.66
0.59±0.02 0.58±0.018
0.59±0.016
0.61±0.018a 0.77±0.009b 0.76±0.029b
0.72±0.012b
Lysozyme was a member of innate humeral factors that elaborated from polymorph nuclear
and mononuclear cells (Moore et al., 2006). Allicin fed chicks in group C showed significant
increase in lysozyme activity at 24 hours post 2nd vaccination with NDV while that in group
B and C showed significant increase at 24 hours post 2nd vaccination of IBDV and at the end
of the experiment compared with control group (Table 5).The present observation is in
accordance with Tatara et al., (2008) and Talpur and Ikhwanuddin (2012), the increase in
lysozyme activity may be due to activation of phagocytic macrophage by allicin.
Nitric oxide is generated during immune and inflammatory response, it is involved in innate
immunity as a toxic agent towards infectious organisms and can induce or regulate death and
function of host immune cells (Coliman, 2001). Chicks fed on different concentrations of
allicin groups B and C revealed significant increase (p<0.05) at 24 hours post 1st dose of
IBDVand 24 hours post 2nd dose of both NDV and IBDV in comparison with control group
(Table 5). Morihara et al (2002) and Feng et al., (2012) showed that allicin treatment
enhanced the production of pro-inflammatory mediators such as IFN, TNFγ, and NO. Also
Kanget et al., (2001) recorded increase in NO production in allicin treated murine peritoneal
macrophages. It was known from the previous study that NO is produced at high levels by
macrophages through its activation Aouatef et al., (2002)
Table 5. Effect of the dietary supplementation of allicin on Serum lysozyme (µg/ml) and nitric oxide
(µmol/ml) of broiler chickens.
PARAMETERS
Time
Group A
(control)
Lysozyme
Group B
(Allicin 125
gm)
Group C
(Allicin
250gm)
Group A
control
1st day post 1st vac
13.79±1.63 16.29±1.92 20.08±1.96
NDV
12.88±.39
st
1 day post 1st vac IBD 18.67±0.84 36.47±0.66 28.57±1.72 11.90±.42a
1st day pos 2nd vac NDV 20.91±1.40a 22.32±1.40 51.58±5.02b 12.71±0.96a
1st day post2nd vac IBD 27.53±2.42a 48.20±3.41b 55.59±3.38b 12.19±0.55a
At end of experament
45.92±3.92a 69.69±6.97b 88.54±5.77b 7.79±0.20a
LSD
23.51
3.71
Means with different small letters are significant different between groups
Nitric oxide
Group B
Allicin 125
gm
Group C
Allicin 250
gm
15.64±.46
19.03±.41b
16.87±0.5b
16.20±1.15b
8.27±0.24b
15.86±.40
18.01±.33b
19.13±0.62b
16.62±0.57b
8.22±0.39b
In regard to humeral immune response, allicin treated groups showed slightly increase in HI
antibody titers for NDV vaccine and ELISA antibody (OD) for IBDV vaccine comparing
with that of control group figures 1 and 2. Our results are coincident with that of Gabor et al.,
(1998) and Haq et al., (1999) who showed that higher garlic supplement increased the
antibodies titer against NDV vaccine, however present data are in contrast to what reported
by Jafari et al., (2008) and Pourali et al., (2010) who showed that adding of garlic powder to
the diet of broilers had no beneficial effect on humoral immune response. This discrepancy
between our results and others is probably due to the use of purified active component
(allicin) in our study while the others used whole garlic extract.
Glutathione is the most abundant low molecular weight thiol compound in cells and plays an
important role in antioxidant defence and detoxification. Glutathione depletion compromises
cell defences against oxidative damage and may lead to cell death (Masella et al., 2005).
Malondialdehyde ( MDA) is the end product of the lipid peroxidation process in which the
peroxyl radicals changed to endoperoxides. The balance between the production of
peroxidants and the scavenging of those oxidants by antioxidants determines the extent of
lipid peroxidation. In the present study , the activities of MDA were reduced while that of
GSH were elevated by allicin treatement in groups B and C in comparison with control group
as shown in (table 6). Allicin (major component of garlic organosulfur) was shown to possess
significant antioxidant activity, which attributed to its rapid action with thiol containing
proteins (Okada et al.,2005; Lee-larungrayub et al., 2006 and Chung, L.Y., (2006). It is
known that garlic increases protein synthesis in damaged tissues and leads to improvement in
the functional status of the cell( Hussein ,2003). Shakiba et al., (2009) recorded that garlic
can scavenge free radicals and prevent GSH depletion. Sharma et al (2010) showed that A.
sativum application led to decrease in lipid peroxidation and increase in the cellular
antioxidant enzyme.
Figure 1. HI titer of NDV vaccine.
Figure 2. ELISA optical densities of IBDV vaccine.
1.2
1.1
ELISA (OD)
1
0.9
control
0.8
low
0.7
high
0.6
0.5
0.4
1st w post
1st v
2nd w post
1st v
1st w post
2nd v
weeks post 1st & 2nd IBDV vaccine
Table 6. Effect of the dietary supplementation of
Allicin on Glutathione and Malondialdehyde
(n.mol/ml)
PARAMETERS Gluathion Malondialdehyde
GROUPS
Group 1
3.81±0.09a 11.91±0.43a
Group 2
4.45±0.07b 10.33±0.19b
Group 3
4.35±0.06b 10.30±0.23b
LSD
0.53
1.58
Means with different small letters are significant
different between groups
Carcass characteristics
Carcass weights, dressing percent, carcass cuts and some organs weight and length
investigated were not affected significantly by allicin supplementation (Table 7). Similar
effect of allicin in the dressing percentages was observed. While the abdominal fat weight
was reduced significantly (p<0.05) by increasing the dose of allicin used. Those finding are
in agreement with those reported by Sharma et al (2010) and Okada et al., (2005).
Table7. Effect of allicin on carcass weight, cuts, dressing percent, relative selected
organs and abdominal fat of broiler chickens*
Parameters (g)
Group A
Group B
Group C
Live wt
1981.5
2011.0
2010.7
Carcass wt
1416.2
1482.5
1481.5
Dressing %
71.5
73.7
73.7
a
b
Abdomen fat
2.21
1.90
1.69c
Head
2.15
2.10
2.22
Neck
5.5
4.9
5.3
Wing
7.9
7.7
7.8
Back
15.6
15.9
16.7
Thigh
19.7
20.6
19.9
Breast
22.0
23.6
23.3
Heart
2.6
2.8
2.5
Liver
4.3
4.0
4.5
Gizzard
4.3
4.2
3.89
Feet
4.5
4.5
4.5
Cecum
1.2
1.1
1.3
Small intestine weight
8.5
8.69
9.1
Small intestine length (cm)
165
175
173
Cecum length (cm)
20.3
21.2
21.0
*Means represent 10 chickens per treatment
Values in the same row with different superscripts are statistically significantly different
(p<0.05).
In conclusion, allicin treatment could enhance broiler chickens growth performance through
its antibacterial and antioxidant effects, decreases abdominal fat deposition and has a positive
impact on innate immunity as it significantly increased phagocytic activity of Peripheral
blood mononuclear cells augment humoral immune response against NDV and IBDV
vaccines. In addition ,allicin treatment maintained oxidant/antioxidant balance.
References
AMAGASE, H., B.L. PETESCH, H. MATSUURA, S. KASUGA, Y. ITAKURA, 2001:
Intake of garlic and its bioactive components. Journal of Nutrition, 131, 955S-962S.
AOUATEF D. EUGENE, M. NELLY, B. YVES, V. Q. PASCALE, 2002: Similar pattern of
expression, No production and cytokine response in genetic and vaccination- acquired
resistance to Marek,s disease. Vet. Imm.&Immunopatho., 85:63-75.
ARI, M. M., R. E. BARDE, D M. OGAH, Y.I. AGADE, N.D. YUSUF, I.D. HASSAN, M.M.
MUHAMMED, 2012: Utilization of garlic (Allium sativum) as a Supplemenary phytogenic
feed additive for broilers fed commercial feeds. Egypt. Poult. Sci. Vol (32) (I): (13-21)
BANERJEE, SK, P.K. MUKHERJEE, S.K. MAULIK, 2001: "Garlic as an Antioxidant: The
Good, The Bad and The Ugly". Phytotherapy Research 17 (2): 97–106.
BAUTISTA D.M., P. MOVAHED, A. HINMAN, H.E. AXELSSON, O. STERNER, E.D.
HOGESTATT, D. JULIUS, S.E. JORDT, P.M. ZYGMUNT, 2005: "Pungent products from
garlic activate the sensory ion channel TRPA1". Proc Natl Acad Sci USA 102 (34): 12248–
52.
BEARD, C.W., 1989: serological procedures. In: Laboratory manual for the isolation and
identification of avian pathogens published by the American Association of avian
pathologists, 3rd ED., pp: 192-200.
BLOCK E, A.J. DANE, S. THOMAS, R.B. CODY, 2010: "Applications of Direct Analysis
in Real Time–Mass Spectrometry (DART-MS) in Allium Chemistry. 2-Propenesulfenic and
2-Propenesulfinic Acids, Diallyl Trisulfane S-Oxide and Other Reactive Sulfur Compounds
from Crushed Garlic and Other Alliums". Journal of Agricultural and Food Chemistry 58 (8):
4617–25.
BOS, H., W.D.E. SOUZA, 2000: Phagocytosis of yeast: a method for concurrent
quantification of binding and internalization using differential interference contrast
microscopy. Journal of Immunolo Methods,238:29-43.
CAVALLITO, C.J., J.S. BUCK, C.M. SUTER, 1994: Allicin, the antibacterial principle of
Allium sativum. Determina-tion of the chemical composition. Journal of the Ameri-can
Chemical Society, 60, 1952-1958.
CHO, SJ., DK. RHEE., S. PYO, 2006: Allicin, a major component of garlic , inhibits
apoptosis of macrophage in a depleted nutritional state. Nutrition. 22(11-12):1177-84.
CHOUDHARY, R., 2008: Benificial effect of Allium sativum and Allium tuberosumon
experimental hyperlipidemia and atherosclerosis. Pak. J. Physiol. 4, 7–9.
CHUNG, L.Y., 2006: The antioxidant properties of garlic compounds: allylcysteine, alliin,
allicin, and allyl disulfide. J. Med. Food. 9, 205–213.
COLEMAN, J.W., 2001: Nitric oxide in immunity and inflammation. International
Immunopharmacology, 1:1397-1406.
DEMIR, E., S. SARICA, M.A. OZCAN, M. SUICMEZ, 2003: The use of natural feed
additives as alternatives for an an-tibiotic growth promoter in broiler diets. British Poultry
Science, 44, S44-S45.
ELLMAN, G.L., 1959: Tissue sulfdrylgroups.ArchBiochem.Biophys,82:70-77.
ERIC BLOCK, 1985:"The chemistry of garlic and onions". Scientific American 252 (March):
114–9.
FENG Y., X. ZHU, Q. WANG, Y. JIANG, H. SHANG, L. CUI, Y. CAO, 2012:
"Allicininhanceshost pro-inflammatory immune responses and protects agains acute murine
malaria infection" Malaria journal, 11:26
GABOR, S., P. VILMOS, N. BELA, E. ISTVANNE, N. GYORGYET, 1998: New type of
immune-stimulant to increase antibody production in response to viral and bacterial vaccines.
Magyar AllatorvosokLapja, 120:719-721.
HAQ, A., K.A. MERAJ, S. RASOOL, 1999: Effect of supplementing A. sativum (Garlic)
andAzadirachtaindica (Neem) leaves on broiler feeds on their blood cholesterol triglycerides
and antibody titer. INT. J. Agri. Biol., 125-127.
HORTON, G.M.J., M.J. FENNEL, B.M. PRASAD, 1991: Effects of dietary garlic on
performance, carcass composition and blood chemistry changes in broiler chickens. Can. J.
Anim. Sci., 71: 939-942.
HUSSEIN, A.A., 2003: Mechanism of action of the Egyptian garlic ‘‘Allium sativum” on
excitable tissues. Pak. J. Bio. Sci. 6 (8), 782–788.
IJI, P.A., A. SAKI, D.R. TIVEY, 2001: Body and intes-tinal growth of broiler chicks on a
commercial starter diet. 1. Intestinal weight and mucosal development. British Poultry
Science, 42, 505-513.
JAFARI R.A., M. GHORBANPOOR, S. HOSHMANDDIARJAN, 2008: Effect of dietary
garlic on serum antibody titer against Newcastle disease vaccine in broiler chicks.Journal of
Biological Sciences 8(7):1258-1260.
JAVANDEL, F., B. NAVIDSHAD, J. SEIFAVATI, G.H. POURRAHIMI, S.
BANIYAGHOUB, 2008: The favourite dosage of garlic meal as a feed additive in broiler
chickens rations. Pak. J. Biol. Sci., 11 (13): 1746-1749.
KANGET, N.S, E.Y. MOON, C.G. CHO, S. PYO, 2001: Immunomodulatingeffect of garlic
component, allicin, on murine peritoneal macrophages, Nutrition Research 21 (2001) 617–
626
LEE-LARUNGRAYUB, N., V. RATTANAPANONE, N. CHANARAT, J.M. GEBICKI,
2006: Quantitative evaluation of the antioxidant properties of garlic and shallot preparations.
J. Nutr. 22, 266–274.
LEWIS, M.R., S.P. ROSE, A.M. MACKENZIE, L.A. TUKER, 2003: Effect of plant extracts
on performance of male broiler chickens. Br. Poult. Sci., 44 (1): 43-44.
LINDSEY J. MACPHERSON, BERNHARD H. GEIERSTANGER, VEENA
VISWANATH, MICHAEL BANDELL, SAMER R. EID, SUNWOOK HWANG, ARDEM
PATAPOUTIAN 2005: "The pungency of garlic: Activation of TRPA1 and TRPV1 in
response to allicin]". Current Biology 15 (10): 929–934.
MASELLA, R., BENEDETTO R.D., R. VARI, C. FILESI, C. GIOVANNINI, 2005:
Novelmechanisms of natural antioxidant compounds in biological systems:involvement of
glutathione and glutathione-related enzymes. J. Nutr.Biochem. 16, 577–586.
MOORE, M.M., J.I. ALLEN, A. MCVEIGH, 2006: Environmental prognostic: an integrated
modal supporting lysosomal stress as predictive biomarkers of animal health status "Marine
Environmental Research,61:278-304.
MORIHARA N, I. SUMIOKA, T. MORIGUCHI, N. UDA, E. KYO 2002: Aged garlic
extract enhancesproduction of nitric oxide. Life Sci, 71:509–517
OHKAWA, H., N. OHISHI, K. YAGI, 1979: Assay for lipid peroxides in animals tissues by
thiobarbituric acid reaction. Anal Biochem, 95:351-358.
OKADA, Y., K. TANAKA, I. FUJITA, E. SATO, H. OKAJIMA, 2005: Antioxidant activity
of thiosulfinates derived from garlic. Redox. Rep. 10, 96–102
ONIBI, G.H., B. NAVIDSHAD, J. SEIFAVATI, 2009: Response of broiler chickens in
terms of performance and meat quality to garlic supplementation. African J. Agricultural
Res., 4 (5): 511-517.
PATYA. M, M.A. ZAHALKA, A. VANICHKIN, A. RABINKOV, T. MIRON, D.
MIRELMAN, M. WILCHEK, H.M. LANDER, A. NOVOGRODSKY, 2004: Allicin
stimulates lymphocytes and elicits an anti tumor effect : a possible role of p21 ras"
International immunology. Vol 16 No. 2 ,pp 275-281
PEETERS T.L., G.R. VANTRAPPEN, 1977: Factors influencing lysozyme determination by
lysoplate method. Cli. Chim. Acta ,74:217-255.
PETRIE, A., P. WATSON, 1999: Statistics for Veterinary and Animal Science. 1ST Ed., PP.
90-99, the Blackwell science Ltd, United Kingdom.
POURALI M., S.A. MIRGHELENJ, H. KERMANSHAHI, 2010: Effects of Garlic powder
on productive performance and immune response of broiler chickens challenged with
Newcastle disease virus. Global Veterinaria 4(6):616-621.
QURESHI, A.A., N. ABUIRMEILEH, Z. DIN, C.E. ELSON, W.C. BURGER, 1983:
Inhibition of cholesterol and fatty acid biosynthesis in liver enzymes and chicken hepatocytes
by polar fractions of garlic. Lipids, 18, 343- 348.
SALMAN, H., M. BERGMAN, H. BESSLER, I.D.J. PUNSKY, H. ALDETTI, H. 1999:
Effect of a garlic derivative (alliin) on peripheral blood cellimmune responses. International
Journal of Immunopharmacology 21 589-59
SHAKIBA, Y., A. MOSTAFAIE, D. ARSHADI, B. SABAYAN, 2009: Application of
garlicorganosulfur compounds in prevention of cyclosporine A-induced hepatotoxicity. Irn.J.
Med. Hypotheses Ideas. 3 (3), 78
SHARMA V., A. SHARMA, L. KANSAL, 2010: The effect of oral administration of
extracts on lead nitrate induced toxicity in male mice " food and chemical toxicology 48,
928-936
SELA U., S. GANOR, I. HECHT, A. BRILL, T. MIRON, A. RABINKOV, M. WILCHEK,
D. MIRELMAN, O. LIDER, R. HERSHKOVIZ 2004: "Allicin inhibits SDF-1alpha-induced
T cell interactions with fibronectin and endothelial cells by down-regulating cytoskeleton
rearrangement, Pyk-2 phosphorylation and VLA-4 expression". Immunology 111 (4): 391–
399.
TALPUR D.A., M. IKHWANUDDIN 2012: Dietary effect of garlic (Alliumsativum) on
haemato-immunological parameters, survival, growth, and disease resistance
againstVibrioharveyiinfection in Asian sea bass, Latescalcarifer ( Bloch)
TATARA, M.R., E. SLIWA, K. DUDEK, A. GAWRON, T. PIERSIAK, 2008: Aged garlic
extract and allicin improve performance and gastrointestinal tract development of piglets in
artificial sow. Ann. Agric.Environ.Med., 15:63-69.
TOLLBA, A.A.H., M.S.H. HASSAN, 2003: Using some natural additives to improve
physiological and productive performance of broiler chicks under high temperature
conditions. Black cumin (niglla sativa) or Garlic (allium sativum). Poultry Science, 23, 327340.
VAIDYA VIPRAJA, U. KEITH INGOLD, DEREK A. PRATT, 2009: "Garlic: Source of the
Ultimate Antioxidants – Sulfenic Acids". Angewandte Chemie 121 (1): 163–6.
YANG,M.D., M. B. RAZAASIM, XIAOGANG JIANG, M. D. BO ZHONG, M. D.
MUHAMMAD SHAHZAD, FUJUN ZHANG, M.D. LU SHEMIN, 2010: Nitric oxide in
both bronchoalveolar lavage fluid and serum is associated with pathogenesis and severity of
antigen-induced pulmonary inflammation in rats .Journal Of Asthma, 47:135-14