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Received: 1 December 2020    Revised: 31 January 2021    Accepted: 19 February 2021

DOI: 10.1111/asj.13540

ORIGINAL ARTICLE

Evaluation of the protective effect of lycopene on growth

performance, intestinal morphology, and digestive enzyme
activities of aflatoxinB1 challenged broilers

Md Touhiduzzaman Sarker  | Zhi Yue Wang | Haiming Yang | Xiaoli Wan |

Asare Emmanuel

College of Animal Science and Technology,

Yangzhou University, Yangzhou City, China Abstract
The current study was conducted to investigate the protective efficiency of dietary
Correspondence
Zhi Yue Wang, Department of Animal lycopene (LYC) supplementation on growth performance, intestinal morphology, and
Nutrition and Feed Science, College of digestive enzyme activities aflatoxinB1 (AFB1) challenged broilers. A total of 240 days
Animal Science and Technology, Yangzhou
University, Wenhui East Road 78# Yangzhou old Arber across male broiler chicks were randomly allocated in five treatments and
city, Jiangsu province 225009, China. six replicates (eight birds per replicate); feed and water were provided ad libitum
Email: [email protected]
during the 42 days experiment. The treatment diets were as follows: (i) Basal diet
Funding information (control), (ii) Basal diet + 100 µg/kg AFB1 contaminated diet, (iii) Basal diet + 100 µg/
Data Center of Management Science,
National Natural Science Foundation of kg AFB1 + 100 mg/kg LYC1, (iv) Basal diet + 100 µg/kg AFB1 + 200 mg/kg LYC2,
China -­Peking University, Grant/Award and (v) Basal diet + 100 µg/kg AFB1 + 400 mg/kg LYC3. The results showed that the
Number: 31802095
addition of LYC to AFB1 contaminated broiler diets significantly increased (p < .05)
average daily gain (ADG) and decreased feed conversion ratio (FCR) compared to the
AFB1 diet. AFB1 diet decreased the intestinal villus height (VH) and crypt depth ratio
(VCR) while increasing the crypt depth (CD). However, dietary LYC supplemented
diets relieved the intestinal morphological alterations. Dietary LYC supplementation
(200 and 400 mg/kg) significantly improved (p  < .05) intestinal digestive enzyme
amylase and lipase activities with AFB1 contaminated diet. These findings suggested
that LYC is a promising feed supplement in the broiler industry, alleviating the harm-
ful effects of AFB1.

KEYWORDS

aflatoxinB1, broiler, growth performance, intestine, lycopene

1 |  I NTRO D U C TI O N increased susceptibility to diseases in several animal species (Peng

et al., 2016; Shivachandra et al., 2003; Yunus et al., 2011). Previous
Mycotoxins are natural compounds produced by fungi, and afla- researches have shown that poultry was extremely sensitive to the
toxinB1 (AFB1) is the most commonly known type of mycotoxins, toxic and carcinogenic action of AFB1, resulting in millions of dollars
which are mainly produced by the Aspergillus flavus and Aspergillus in annual losses to farmers due to reduced growth rate, eggs and
paraciticus (Diaz et al., 2003). AFB1 is considered the most highly meat production, increased susceptibility to disease, and other ad-
widespread and toxic type of aflatoxin (Alpsoy & Yalvac, 2011). The verse effects (Guo et al., 2012; Rawal et al., 2010; Shi et al., 2012;
negative effects of AFB1 have also been well documented, includ- Yunus et al., 2011). Feeding naturally AFB1 contaminated diets in
ing reduced performance, decreased immune system function, and the starter and grower stage compromised growth performance,

Anim Sci J. 2021;92:e13540. wileyonlinelibrary.com/journal/asj |

© 2021 Japanese Society of Animal Science     1 of 9
https://doi.org/10.1111/asj.13540
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2 of 9       SARKER et al.

intestinal morphology, changes in digestive physiology, and devel- 2 | M ATE R I A L S A N D M E TH O DS

opment in ducks (Feng et al., 2017).
Recently, there has been an increasing awareness of the adverse This experiment was conducted in the Animal Nutrition and Feed
effects of AFB1 on vulnerable structures in the intestine and the im- Science Department, College of Animal Science and Technology,
pairment of intestinal integrity (Bouhet & Oswald, 2005). The intes- Yangzhou University, China, and all protocols were approved by
tinal tract is the first target organ which is been destructed by the Yangzhou University animal care and use committee.
activities of AFB1 when it enters into the body of the animals, thus,
compared with other organs (liver, lungs, spleen, and kidney). This
toxin exerts greater impacts on the small intestine. Additionally, the 2.1 | Experimental birds, diets, and husbandry
animal's intestinal health status depends on the integrity and inflam-
matory levels of the animals' intestine (Bischoff, 2011). It was shown A total of 240 one-­d ay-­o ld male Arbor Acres broilers were ob-
that like other mycotoxins, AFB1 altered the intestinal absorbing bar- tained from a commercial hatchery (Nantong, Jiangsu, China).
rier (Ali Rajput et al., 2017), reduced the activity of digestive enzymes, The broiler chicks were randomly assigned to five treatments
and disturbed the energy metabolism of the intestinal cells as well with six replicates (eight birds in each pen). Treatments were as
as the gluconeogenesis, fatty acids synthesis (Ling et al., 2016; Long follows: (a) A basal diet containing neither AFB1 nor LYC (Control)
et al., 2016; Pinton & Oswald, 2014). Digestive enzymes such as am- and (b) a basal diet containing 100 µg/kg AFB1 . The other three
ylase and lipase are secreted by the intestinal tract; they are closely treatment groups were supplemented with LYC at 100, 200, and
related to digestion and absorption of nutrients as well as growth and 400 mg/kg on the 100 µg/kg AFB1 . All broilers were raised in
development. They also play vital tasks such as muscle building, de- the cage breeding system in the environmentally controlled room
stroying toxins, and breakdown food partials for nutrient absorption. during the whole experimental period. The room temperature
Lycopene (LYC), a naturally occurring carotenoid found in was 32°C–­3 4°C for the first 3 days, then gradually decreased to
tomatoes and tomato products, has drawn considerable atten- 22  ± 1°C by 2°C–­3 °C each week and maintained until 42 days
tion as a potential chemopreventive agent (Boeira et al., 2014). experimental period. All broilers in the experimental period free
It was one of the most effective antioxidants in the carotenoid access to feed and water with a lighting cycle of 23 hr light and
family, and its activity against biological reactive oxygen species 1 hr dark. Corn-­s oybean-­b ased diets were formulated accord-
may prevent or ameliorate oxidative damage to cells and tissues ing to NRC (1994) to meet the nutrient requirements from 1 to
(Boeira et al., 2014; Yonar, 2012). It was known to exert an effec- 21 days (starter) and 22–­42 days (grower) experimental periods
tive free radical scavenging activity, and this action could be ben- for broilers (Table 1).
eficial to poultry production because free radicals were formed AflatoxinB1 (Purity ≥98%, HPLC) and LYC (Purity ≥80%, HPLC)
under stress, fast-­g rowing, high production rates, and intensive were purchased from Shanghai Yuanye Biotechnology Co. Ltd.
metabolic conditions of poultry (Englmaierová et al., 2011). In Previous studies have demonstrated that 100 µg/kg AFB1 in diets
recent years, LYC, as a beneficial carotenoid, has received con- had obvious adverse effects on broilers, and appropriate levels of
siderable scientific attention (Sahin et al., 2006, 2014). Dietary LYC supplementation in the diet (200–­4 00 mg/kg) could provide op-
supplementation with LYC has also been associated with lower timal protective effects against heat stress broilers (Liu et al., 2016;
risks of cancers, cardiovascular diseases, and other humans and Peng et al., 2014; Sahin et al., 2016). Based on this information, an
animals' disorders (Mordente et al., 2011). Moreover, dietary LYC appropriate toxic concentration and dietary supplementation levels
supplementation is known to protect the animal intestine (struc- of LYC were chosen.
ture and tissue) from damage after coming into contact with
harmful agents like pathogen, toxins, or any other foreign anti-
gen. Mumtaz et al. (2013) demonstrated that supplementation of 2.2 | Samples collection and measurements
LYC with vitamins A, C, and E to radiation exposed rats' intestinal
toxicity could protect against intestinal damage. Previous studies The broiler weight and feed intake (FI) were recorded at 21 and
(Saada et al., 2010; Yilmaz & Kaya, 2019; Yucel et al., 2016) have 42 days for calculation of average daily gain (ADG) and average daily
demonstrated the beneficial effects of dietary supplementation feed intake (ADFI) and feed conversion ratio (FCR). At the end of
of LYC in the AFB1 contaminated diet on the improved intestinal the experiment, the broilers were euthanized by cervical dislocation
structure of animals. However, the protective effects of dietary and the duodenum, jejunum, and ileum were removed. The sample
LYC in the intestine damage on growth performance against of small intestine tissues (approximately 2 cm from the middle of
AFB1 challenged broilers have not been reported. Therefore, this duodenum, jejunum, and ileum, respectively) was collected and then
study aimed to evaluate the protective efficiency of different stored in 10% formaldehyde solution for further analysis. Intestinal
levels of dietary LYC supplementation on growth performance, digesta (duodenum, jejunum, and ileum) were collected into a plastic
intestinal morphology, and digestive enzyme activities in AFB1 tube to determine digestive enzyme activities, and all samples were
challenged broilers. stored at −40°C until analysis.
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TA B L E 1   Composition of feed ingredients (g/kg) and nutrient and stained with hematoxylin and eosin (H&E). The values of villus
level (%) height (VH), crypt depth (CD), and ratio (VCR) on every segment
Items 1–­21 days 22–­42 days were determined using a light microscope (Nikon YS100) attached
to a computer.
Ingredients (g/kg)
Corn 57.10 61.00
Soybean meal 31.00 28.00
2.4.1 | Intestinal enzyme activity assay
Corn gluten meal 4.00 1.60
Soybean oil 3.00 1.30 Trypsin (catalog No. A080-­2), amylase (catalog No. C016-­1), and li-
Dicalcium phosphate 2.00 2.40 pase (catalog No. A054-­1-­1) activities were measured by using cor-
Limestone 1.20 4.00 responding commercial kits purchased from Nanjing Jian Cheng
L-­Lysine 0.20 0.25 Bioengineering institution. The measurements were performed ac-
DL-­Methionine 0.20 0.15 cording to the kit's instructions.
a
Premix   0.31 1.00
Sodium chloride 0.30 0.30
Calculated nutrient levels (%)
2.5 | Statistical analysis
Metabolizable energy (MJ/kg) 12.61 12.96
Statistical analysis of all data was performed using SPSS software
Crude protein 21.36 19.44
(Version 21; SPSS Inc.). One-­way ANOVA with a post hoc test was
Calcium 1.00 0.93
used to elucidate significant differences. Duncan's test was used for
Available phosphorus 0.46 0.39
multiple comparisons when a significant difference was detected.
Lysine 1.09 1.05 Differences were considered to be statistically significant at p < .05,
Methionine 0.56 0.47 and .05 < p <.10 was considered to be a trend towards significant. All
Arginine 1.27 1.16 data were presented as the mean and standard error of the means
Methionine + cysteine 0.91 0.80 (SEM).
a
The premix provided per kilogram of diet: vitamin A (retinyl acetate),
12,000 IU; vitamin D3 (cholecalciferol), 2,500 IU; vitamin E (DL-­
α-­tocopheryl acetate), 20 IU; menadione, 1.3 mg; thiamin, 2.2 mg; 3 | R E S U LT S
riboflavin, 8.0 mg; nicotinamide, 40 mg; choline chloride, 400 mg;
calcium pantothenate, 10 mg; pyridoxine HCl, 4 mg; biotin, 0.04 mg;
folic acid, 1.0 mg; vitamin B12 (cobalamin), 0.013 mg; Fe (from ferrous 3.1 | Growth performance
sulfate), 80 mg; Cu (from copper sulphate), 8.0 mg; Mn (from manganese
sulphate), 110 mg; Zn( from zinc sulfate), 60 mg; I (from calcium iodate), Data from Table 2 represent the effects of dietary treatment on
1.1 mg; Se (from sodium selenite), 0.3 mg.
broiler growth performance. No significant effect (p < .05) was ob-
served in ADFI throughout the experimental period. However, broil-
ers exposed to AFB1 diet had decreased (p < .05) ADG and increased
2.3 | Preparation of intestinal digesta homogenate FCR during the experimental trial compared to the control group.
Broilers fed diet AFB1 + LYC1 improved (p = .007) ADG as compared
Approximately 0.2 g of intestinal digesta samples (duodenum, jeju- to AFB1 at 1–­21 days. Broilers fed diet AFB1 + LYC2 and AFB1 + LYC3
num, and ileum digesta) was homogenized with cold physiological significantly improved (p < .05) the ADG in 22–­42 and 1–­42 days.
saline solution (4°C) at a ratio of 1:9 (wt/vol) using an ultraturrax Additionally, a trend of decreased (p  = .07) in FCR was ob-
homogenizer (Shanghai Jing Xin). The homogenized solution was served in AFB1 + LYC2 group at 22–­42 days while AFB1 + LYC2, and
then centrifuged (cence DL-­5M) at 2,500 rpm for 10 min at 4°C, and AFB1 + LYC3 groups decreased (p < .05) the FCR in 1–­42 days com-
the supernatant was obtained and immediately stored at −20°C for pared to AFB1 group.
further analysis.

3.2 | Intestine morphology
2.4 | Intestinal morphology
Comparatively, data on duodenal morphology indicated that AFB1
The fixed small intestine segments (duodenum, jejunum, and significantly decreased the VH and VCR, while CD was significantly
ileum) were soaked through a graded series of ethanol and xylene, increased (p < .05) compared to the control. However, dietary LYC
embedded in paraffin, sections at 5 µm with a Leica RM 2016 mi- supplementation increased VH and VCR while decreasing (p < .05)
crotome (Leica Biosystems Inc.). The sections were deparafinizied CD as compared to the AFB1 group on 21 and 42 days (Table 3 and
with xylene and rehydrated through graded dilutions of ethanol Figure 1). Additionally, on 21 days, broilers fed diet AFB1 jejunal
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TA B L E 2   Effects of LYC on growth performance of broilers fed diets contaminated with AFB1

Dietary treatments

Parameters Control AFB1 AFB1 + LYC1 AFB1 + LYC2 AFB1 + LYC3 SEM p-­value

1–­21  days
ADG (g/day) 32.55a 28.54c 30.76bab 30.47bc 30.35bc 0.364 .007
ADFI (g/day) 48.25 44.09 46.91 45.90 45.77 0.512 .118
FCR (Feed/gain) 1.48b 1.54a 1.52ab 1.50ab 1.50ab 0.006 .039
22–­42  days
ADG (g/day) 70.27a 63.03b 64.48b 68.41a 67.53a 0.648 <.001
ADFI (g/day) 136.07 127.38 128.07 133.70 132.35 1.388 .222
FCR (Feed/gain) 1.93b 2.01a 1.98ab 1.95b 1.96ab 0.010 .074
1–­42  days
ADG (g/day) 51.51a 46.00 c 47.32bc 49.44ab 48.59b 0.492 <.001
ADFI (g/day) 92.37 86.09 86.90 89.64 88.38 0.907 .202
FCR (Feed/gain) 1.78b 1.87a 1.83ab 1.81b 1.81b 0.007 .016

Note: Means with different superscripts (a, b, c) in the same row were significantly different at p < .05, and .05 < p < .10 was considered to be a
tendency towards significant. SEM, pooled standard error of the means (n = 6).
Abbreviations: ADFI, average daily feed intake; ADG, average daily gain; AFB1 +  LYC1-­(100  µg/kg AFB1 + 100 mg/kg LYC); AFB1 + LYC2 (100 µg/kg
AFB1 + 200 mg/kg LYC); AFB1 + LYC3 (100 µg/kg AFB1 + 400 mg/kg LYC); AFB1-­(100  µg/kg AFB1); AFB1, aflatoxinB1; FCR, feed conversion ratio;
LYC, lycopene; SEM, standard error of the mean.

TA B L E 3   Effects of LYC on duodenum morphology of broiler fed with AFB1 contaminated diets at 21 and 42 days

Dietary treatments

Parameters Days Control AFB1 AFB1 + LYC1 AFB1 + LYC2 AFB1 + LYC3 SEM p-­value

a c b ab ab
VH (µm) 21 1,415.75 1,136.01 1,278.17 1,311.32 1,317.35 24.605 .003
42 1801.66a 1529.98c 1645.82b 1673.89b 1,670.21b 20.157 <.001
CD (µm) 21 205.82b 240.67a 217.18b 213.87b 201.85b 3.900 .007
b a b b
42 266.45 320.74 271.00 259.21 254.66b 6.828 .007
VCR (µm/µm) 21 6.91a 4.73c 5.89b 6.17b 6.05ab 0.170 <.001
a b a a
42 6.82 4.81 6.14 6.71 6.69a 0.169 <.001

Note: Means with different superscripts (a, b, c) in the same row were significantly different at p < .05, and .05 < p < .10 was considered to be a
tendency towards significant. SEM, pooled standard error of the means (n = 6).
Abbreviations: AFB1 + LYC1 (100 µg/kg AFB1 + 100 mg/kg LYC); AFB1 +  LYC2-­(100  µg/kg AFB1 + 200 mg/kg LYC); AFB1 +  LYC3-­(100  µg/kg
AFB1 + 400 mg/kg LYC); AFB1-­(100  µg/kg AFB1); AFB1, aflatoxinB1; CD, crypt depth; LYC, lycopene; SEM, standard error of the mean; VCR, villus
height and crypt depth ratio; VH, villus height.

CD was significantly increased while VCR was also significantly de- supplementation increased VH and VCR on 21 and 42 days while de-
creased (p < .05) as compared to the control. creased (p < .05) CD on 21 days compared to AFB1 group (Table 5).
Moreover, on 42 days, fed diet AFB1 recorded a decreased in VH
and VCR compared to the control. On the other hand, irrespective of
the dosage, LYC addition decreased (p = .008) CD while the addition 3.3 | Digestive enzyme activities
AFB1 + LYC2 and AFB1 + LYC3 increased (p = .002) the VH and VCR
at 42 days compared to the AFB1 (Table 4). Similarly, data of ileum Data on duodenum enzyme activities indicated that AFB1 signifi-
morphology on 21 days indicated that broilers fed diet AFB1 signifi- cantly decreased (p < .05) the amylase and lipase activities compared
cantly decreased (p  < .05) VH and VCR while CD was also signifi- to the control at 21 and 42 days. However, dietary LYC supplementa-
cantly increased (p < .05) as compared to the control. Furthermore, tion AFB1 + LYC2 and AFB1 + LYC3 on 21 days, and AFB1 + LYC2 on
on 42 days, fed diet AFB1 recorded a decreased in VH and VCR com- 42 days, significantly increased (p < .05) amylase and lipase activities
pared to the control. Besides, except CD on 42 days, dietary LYC compared to the AFB1 group (Table 6).
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F I G U R E 1   Histological structure of
Hematoxylin and Eosin (H&E) staining
duodenum at 21 and 42 days of age.
Control, AFB1-­(100  μg/kg AFB1),
AFB1 +  LYC1-­(100  μg/kg AFB1 + 100 μg/
kg LYC), AFB1 +  LYC2-­(100  μg/
kg AFB1 + 200 mg/kg LYC),
AFB1 +  LYC3-­(100  μg/kg AFB1 + 400 mg/
kg LYC). Scale bar–­4 0 µm

TA B L E 4   Effects of LYC on jejunum morphology of broiler fed with AFB1 contaminated diets at 21 and 42 days

Dietary treatments

Parameters Days Control AFB1 AFB1 + LYC1 AFB1 + LYC2 AFB1 + LYC3 SEM p-­value

VH (µm) 21 838.17 769.44 835.42 842.32 829.61 11.767 .133

a b b a a
42 1,229.49 1,067.06 1,090.04 1,222.20 1,213.78 15.390 <.001
CD (µm) 21 153.62b 188.63a 168.72b 157.96b 160.12b 3.606 .008
42 212.26 222.26 223.50 222.40 220.99 3.648 .886
VCR (µm/µm) 21 5.48a 4.07b 5.00a 5.32a 5.20a 0.126 <.001
42 5.82a 4.81c 4.95bc 5.56a 5.44ab 0.101 .002

Note: Means with different superscripts (a, b, c) in the same row were significantly different at p < .05, and .05 < p < .10 was considered to be a
tendency towards significant. SEM, pooled standard error of the means (n = 6).
Abbreviations: AFB1 +  LYC1-­(100  µg/kg AFB1 + 100 mg/kg LYC); AFB1 +  LYC2-­(100  µg/kg AFB1 + 200 mg/kg LYC); AFB1 +  LYC3-­(100  µg/kg
AFB1 + 400 mg/kg LYC); AFB1-­(100  µg/kg AFB1); AFB1, aflatoxinB1; CD, crypt depth; LYC, lycopene; SEM, standard error of the mean; VCR, villus
height and crypt depth ratio; VH, villus height.

TA B L E 5   Effects of LYC on ileum morphology of broiler fed with AFB1 contaminated diets at 21 and 42 days

Dietary treatments

Parameters Days Control AFB1 AFB1 + LYC1 AFB1 + LYC2 AFB1 + LYC3 SEM p-­value

a b a a a
VH (µm) 21 535.11 440.09 513.67 538.10 508.52 10.457 .011
42 1,069.73a 934.83b 1,033.29a 1,047.91a 1,033.83a 11.446 .010
CD (µm) 21 114.52b 153.02a 128.85b 114.63b 107.41b 4.208 <.001
42 177.34 209.51 185.00 181.84 180.31 4.404 .128
a b a a a
VCR (µm/µm) 21 4.72 2.90 4.08 4.73 4.72 0.160 <.001
42 6.09a 4.63b 5.70a 5.91a 5.63a 0.129 <.001

Note: Means with different superscripts (a, b, c) in the same row were significantly different at p < .05, and .05 < p < .10 was considered to be a
tendency towards significant. SEM, pooled standard error of the means (n = 6).
Abbreviations: AFB1 +  LYC1-­(100  µg/kgAFB1 + 100 mg/kg LYC); AFB1 +  LYC2-­(100  µg/kg AFB1 + 200 mg/kg LYC); AFB1 +  LYC3-­(100  µg/kg
AFB1 + 400 mg/kg LYC); AFB1-­(100  µg/kg AFB1); AFB1, aflatoxinB1; CD, crypt depth; LYC, lycopene; SEM, standard error of the mean; VCR, villus
height and crypt depth ratio; VH, villus height.

In addition, broilers fed diet AFB1 jejunum amylase and lipase Similarly, ileum lipase activities of broilers fed diet AFB1 on
activities on 21 and 42 days, trypsin activities on 42 days, signifi- 21 days, and amylase and lipase activities on 42 days were signifi-
cantly decreased (p  < .05) compared to the control. On the other cantly decreased (p  < .05) compared to the control. Conversely,
hand, irrespective of the dosage, LYC supplementation AFB1 + LYC2, dietary LYC supplementation AFB1 + LYC2 and AFB1 + LYC3 signifi-
and AFB1 + LYC3 increased (p < .05) amylase and lipase activities on cantly increased (p < .05) amylase activities on 42 days, compared to
42 days compared to the AFB1 group (Table 7). the AFB1 group (Table 8).
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TA B L E 6   Effect of LYC on duodenum digestive enzyme activity of broiler fed with AFB1 contaminated diets at 21 and 42 days

Dietary treatments

Parameters Days Control AFB1 AFB1 + LYC1 AFB1 + LYC2 AFB1 + LYC3 SEM p-­value

Trypsin (U/gprot) 21 2.01 1.49 1.73 1.88 1.78 0.068 .166

42 1.91 1.55 1.81 1.88 1.79 0.063 .437
Amylase (U/gprot) 21 179.51a 135.54c 153.73bc 168.75ab 158.80ab 4.047 .003
a b ab a ab
42 170.91 148.94 155.37 167.52 162.18 2.741 .061
Lipase (U/gprot) 21 3.78a 2.87b 3.26ab 3.66a 3.58a 0.093 .006
a c c ab bc
42 3.59 2.83 2.91b 3.41 3.11 0.074 <.001

Note: Means with different superscripts (a, b, c) in the same row were significantly different at p < .05, and .05 < p < .10 was considered to be a
tendency towards significant. SEM, pooled standard error of the means (n = 6).
Abbreviations: AFB1 +  LYC1-­(100  µg/kgAFB1 + 100 mg/kg LYC); AFB1 +  LYC2-­(100  µg/kg AFB1 + 200 mg/kg LYC); AFB1 +  LYC3-­(100  µg/kg
AFB1 + 400 mg/kg LYC); AFB1-­(100 µg/kg AFB1); AFB1, AflatoxinB1; LYC, Lycopene; SEM, standard error of the mean.

TA B L E 7   Effect of LYC on jejunum digestive enzyme activity of broiler fed with AFB1 contaminated diets at 21 and 42 days

Dietary treatments

Parameters Days Control AFB1 AFB1 + LYC1 AFB1 + LYC2 AFB1 + LYC3 SEM p-­value

Trypsin (U/gprot) 21 1.76 1.46 1.49 1.68 1.65 0.049 .260

a b b ab ab
42 2.23 1.95 1.95 2.17 2.03 0.036 .030
Amylase (U/gprot) 21 177.22a 152.74b 158.96ab 170.99ab 164.92ab 2.966 .063
42 139.00a 113.05c 122.09bc 130.69ab 127.81b 2.072 <.001
a b b ab
Lipase (U/gprot) 21 3.65 2.87 2.98 3.13 3.12ab 0.086 .037
a c c ab ab
42 2.79 1.89 2.07b 2.57 2.44 0.093 .007

Note: Means with different superscripts (a, b, c) in the same row were significantly different at p < .05, and .05 < p < .10 was considered to be a
tendency towards significant. SEM, pooled standard error of the means (n = 6).
Abbreviations: AFB1 +  LYC1-­(100  µg/kg AFB1 + 100 mg/kg LYC); AFB1 +  LYC2-­(100  µg/kg AFB1 + 200 mg/kg LYC); AFB1 +  LYC3-­(100  µg/kg
AFB1 + 400 mg/kg LYC); AFB1-­(100  µg/kg AFB1); AFB1, aflatoxinB1; LYC, lycopene; SEM, standard error of the mean.

4 |  D I S CU S S I O N ADG (Englmaierová et al., 2011), FI (Lira et al., 2010) and could

even alleviate adverse effects of stress conditions on performance
Poultry is an important international food commodity, and the (Sahin et al., 2016). Moreover, LYC supplementation in quail's diets
broiler is the world's most available and major protein source. improved the ADG and FCR even under stress conditions (Sahin
In the recent era, broiler feed is hard to avoid toxic contamina- et al., 2006). In the present study, we were observed that supple-
tion; it is increasingly recognized that long-­term consumption of mentation with different levels of LYC (100, 200, and 400 mg/kg)
AFB1 contaminated feed has detrimental effects on broiler growth to the moldy diets ameliorated the growth suppression induced by
and performance. In this present study, AFB1 (100 µg/kg) signifi- AFB1, as evidenced by significantly increased ADG and decreased
cantly depressed the ADG and increased the FCR but did not in- FCR. So the toxic effect of AFB1 on growth performance can be
fluence the ADFI during the whole experimental period. Similar overcome by the addition of LYC in the broiler diet.
results had been observed in the previous studies, in which AFB1 The intestine acts as feed digestion, absorption function,
(50–­100  µg/kg) could significantly decrease the body weight which also acts as an important fighter against all pathogenic
gain and FCR of broilers, causing economic losses (Bintvihok & bacteria and toxic substances present in the intestinal lumen. The
Kositcharoenkul, 2006; Fan et al., 2013). Another study in ducks intestinal villus and crypt system play an important role in nutri-
fed diets contaminated with 20 µg/kg of AFB1 had a significantly ents absorption and animal growth (Hernandez et al., 2006). These
reduced ADG and increased FCR (Han et al., 2008). The possible roles provide the conditional environment forming the indicators
explanation might be anorexia, listlessness, impairment of general of intestinal health status. It has been suggested that shorter villus
health and the inhibitory effects of AFB1 on protein synthesis and height and deeper crypt depth caused poor nutrient absorption,
lipogeneses. increased secretion of electrolytes and water in the intestinal
Several researches conducted by different authors indicated tract, and compromised animals' growth and performance (Li
that dietary supplemented with LYC in chicken diets increased et al., 2015). In this present study, we found that compared with
SARKER et al. |
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TA B L E 8   Effect of LYC on ileum digestive enzyme activity of broiler fed with AFB1 contaminated diets at 21 and 42 days

Dietary treatments

Parameters Days Control AFB1 AFB1 + LYC1 AFB1 + LYC2 AFB1 + LYC3 SEM p-­value

Trypsin (U/gprot) 21 2.11 1.75 1.93 1.99 1.99 0.077 .721

42 1.89 1.34 1.65 1.86 1.65 0.072 .111
Amylase (U/gprot) 21 154.53 133.74 139.09 150.03 147.94 3.860 .453
a c c ab ab
42 217.33 167.99 188.14b 195.81 193.81 4.510 .070
Lipase (U/gprot) 21 3.75a 2.31b 2.78b 3.03ab 2.93ab 0.141 .017
a b b ab b
42 4.13 3.32 3.39 3.82 3.46 0.097 .028

Note: Means with different superscripts (a, b, c) in the same row were significantly different at p <.05, and .05 < p < .10 was considered to be a
tendency towards significant. SEM, pooled standard error of the means (n = 6).
Abbreviations: AFB1 +  LYC1-­(100  µg/kg AFB1 + 100 mg/kg LYC); AFB1 +  LYC2-­(100  µg/kg AFB1 + 200 mg/kg LYC); AFB1 +  LYC3-­(100  µg/kg
AFB1 + 400 mg/kg LYC); AFB1-­(100  µg/kg AFB1); AFB1, aflatoxinB1; LYC, lycopene; SEM, standard error of the mean.

the control group, the villus height and villus height/crypt depth intestinal structure caused by the AFB1 diet. AFB1 induced distur-
in three parts (duodenum, jejunum, and ileum) of the small intes- bance of digestive enzyme activity and nutrient transporters may
tine were noticeably decreased in AFB1 treated group broilers. lead to intestinal disorders, resulting in the alteration of nutrient
These results were in agreement with previous studies observed digestion, absorption function, and reduced growth performance
by Zhang et al. (2014), who found that 0.3 mg/kg AFB1 decreased of broilers. Matur et al. (2010) found that lipase enzyme activity
jejunum villus height and the villus height/crypt depth ratio and decreased in the duodenum after feeding 0.1 mg/kg AFB1 in Ross
disrupted intestinal barrier cells of the broiler chicken at 21 days female diets.
of age. Similarly, Wan et al. (2013) found that the intestine villus The current study has shown that the addition of different lev-
height and villus height/crypt depth ratio have been decreased els of dietary LYC with AFB1 contaminated diet improved the amy-
when the addition of 100 µg/kg AFB1 as their dietary treatment lase and lipase activities in the small intestine of broilers. A possible
in ducklings. The possible reasons might be that AFB1 reduced explanation for the increased enzyme activities might be ascribed
the nutrient absorption surface area by damaging the broiler in- to the improved intestinal morphology and integrity by dietary LYC
testine structure. The above results indicated that dietary AFB1 supplementation. Also, this result indicated that LYC might have di-
decreased the broiler intestine function by depressing intestinal gestive enhancing properties and thus improved the growth perfor-
morphological development and induced toxic lesions. mance of broilers. Findings of our current study implied that dietary
LYC as carotenoid has anticarcinogenic and anticancer proper- supplementation of LYC alleviated the detrimental effects of AFB1
ties, which primarily responsible for its beneficial effects. Recent on the small intestine, digestive enzyme, and growth performance
studies have found that LYC plays an important role in the preven- of broilers.
tion of gastrointestinal tract cancer. Yucel et al. (2016) found that
LYC was beneficial in protected intestinal damage against metho-
trexate in rats. A study by Itoh et al. (2004) found that LYC pos- 5 | CO N C LU S I O N
sessed a significant radio protective effect on villus and crypts in
the small intestine of abdominally radiated (15 Gy) mice. Data from In conclusion, 100 µg/kg AFB1 in the broiler diet result in some nega-
the current study showed that villus height and villus height/crypt tive effects manifested by a decrease in ADG, an increase in FCR, a
depth ratio of broiler small intestine was significantly increased change in intestinal morphological status, and reduction in digestive
by adding different levels of dietary LYC, thus alleviating AFB1 in- enzyme activities in broilers. Dietary supplementation of different
duced toxicity and improved the histological structure of the small levels (100, 200, and 400 mg/kg) of LYC exhibited beneficial effects
intestine. on growth performance, intestinal morphological status, and diges-
The intestinal enzyme activities have a crucial effect on the an- tive enzyme activities of broiler challenged with AFB1. The results of
imal's physiological processes and life activities (Cho et al., 2012). this study also demonstrated that dietary supplementation of LYC
It is widely believed that the AFB1 application has malabsorption could be incorporated in broiler diets and exhibit promising effects
syndrome regarding macronutrients and reduced activity of diges- to overcome the deleterious effects of AFB1 impact in the broiler
tive enzymes (Devegowda & Murthy, 2005). It has been demon- production.
strated in many studies that AFB1 adversely affected production
performance in the poultry industry (Zaghini et al., 2005). The AC K N OW L E D G M E N T S
present study showed a decrease in the intestinal digestive en- This study was supported by the National Natural Science
zymes, including trypsin, amylase, and lipase in the AFB1 treated Foundation of China (31802095). The assistance of my teachers and
diet. The possible reason might be ascribed to the damage of the lab mates in this work is greatly acknowledged.
 |
8 of 9       SARKER et al.

C O N FL I C T O F I N T E R E S T nutrient digestibility of cherry valley ducks in response to aflatoxin

B1 levels. Livestock Science, 119, 216–­220. https://doi.org/10.1016/j.
The authors declare no conflict of interest.
livsci.2008.04.006
Hernandez, F., Garcia, V., Madrid, J., Orengo, J., Catalá, P., & Megias, M.
ORCID (2006). Effect of formic acid on performance, digestibility, intestinal
Md Touhiduzzaman Sarker  https://orcid.org/0000-0002-0629-7824 histomorphology and plasma metabolite levels of broiler chickens.
British Poultry Science, 47, 50–­ 56. https://doi.org/10.1080/00071​
66050​0 475574
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