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Neurobehavioral and Immune-Histochemical Effects of Daucus
Carota Ethanolic Leaf Extract in Cadmium-Induced Toxicity of the
Hippocampus and Prefrontal Cortex of Adult Wistar Rats
Okechukwu Anyigor-Ogah1, *Chijioke Stanley Anyigor-Ogah2, Clinton Ogbonna Njoku3, Idika Mba
Idika4, Toochukwu Nnamdi Nnama1, Miracle Erinma Chukwuonye2, Agatha Nkechinyere Ekechi2,
Albert N. Eteudo3, Ndudim Ogwuegbu Okezie2,
1Department of Human Anatomy, Alex Ekwueme Federal University, Abakaliki, Ebonyi State, Nigeria
2Department of Family Medicine, Alex Ekwueme Federal University Teaching Hospital, Abakaliki,
Ebonyi State, Nigeria
3Department of Human Anatomy, Ebonyi State University, Abakaliki, Nigeria
4Department of Family Medicine, David Umahi Federal University, Teaching Hospital, Uburu, Ebonyi
State, Nigeria
DOI: https://doi.org/10.51244/IJRSI.2025.120800187
Received: 07 Aug 2025; Accepted: 20 Aug 2025; Published: 18 September 2025
ABSTRACT
Introduction: Chronic exposure to cadmium has been linked to neuro-degenerative disorders due to its ability
to induce neuro-inflammation and apoptosis in critical brain regions such as the hippocampus and prefrontal
cortex.
Objectives: This study evaluated the neurobehavioral and immune-histochemical effects of Daucus
carota ethanolic leaf extract (CLE) in cadmium-induced toxicity of the hippocampus and prefrontal cortex of
adult wistar rats.
Materials and methods: Thirty adult male Wistar rats (weighing 150–180g) were assigned into five groups (6
per group). Group 1 (normal control) received water, Group 2 (Cadmium-only) received cadmium chloride (5
mg/kg). Group 3 received only CLE (400 mg/kg). Groups 4 and 5 received cadmium chloride and CLE at
doses of 200 mg/kg and 400 mg/kg, respectively. Neuro-behavioral tests were conducted to assess cognitive
and emotional responses. Brain tissues were harvested for biochemical analysis as well as immune-
histochemical evaluation of neuronal integrity. Data were analyzed using GraphPad Prism version 8 and
presented as Mean ± SEM. Statistical comparisons were made using one-way ANOVA followed by Tukey’s
post hoc test, with significance set at p < 0.05.
Results: Cadmium exposure significantly impaired cognitive function and triggered neuro-inflammation.
Group C showed weight loss compared to the control group, indicating systemic toxicity. However, CLE
treatment ameliorated these changes in a dose-dependent manner. The Cadmium + CLE (200 mg/kg) and
Cadmium + CLE (400 mg/kg) groups exhibited significant improvements compared to the Cadmium-only
group. The highest dose (400 mg/kg) demonstrated the most pronounced neuroprotective effects, with weight
parameters approaching those of the control group. Histamine level were lowered significantly in group B
when compared to the control group A at p<0.05, acetylcholine level was significantly lowered in all the
treated groups when compared to group B at p<0.05.
Conclusion: The CLE exhibits potent neuroprotective properties against cadmium-induced neurotoxicity,
hence could serve as a promising natural intervention for mitigating heavy metal-induced cognitive and
neuronal impairments.
Key words: Neurobehavioral, Immune-histochemical, Cadmium, Hippocampus, Prefrontal
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INTRODUCTION
Heavy metal toxicity is one of the oldest environmental problems and remains a serious health concern today.1
The toxicity and perceived permanency of heavy metal contaminations in the environment have sparked
growing concern.2 High concentrations of these indestructible and non-biodegradable metals are hazardous to
all forms of life.3 One of these heavy metals known to be part of this is cadmium (Cd).4 Therefore, it is
recognized as an important environmental contaminant.5
The greatest exposure to Cd occurs in the metallurgical industry (in zinc smelters or in units where pig iron is
purified).6 Countries with a significant prevalence of this metal toxicity in humans include Nigeria, India, the
Philippines, Indonesia, Pakistan, Peru, the United States of America, China, Brazil, France, and Mexico.7 The
continued use of Cd in industry drastically affects the environment, resulting in high exposure of humans to the
element.8
Traditional medicine is becoming the mainstay of basic healthcare in developing and poor nations.9
Comparatively speaking, plant-derived medications are more affordable and readily available than their
synthetic equivalents. An example of this medicinal plant is the Carrot (Daucus carota).10 The greatest
nutritional interest in carrots stems from their bioactive content, but research has also focused on carrots as a
source of fiber. Studies have shown that carrot extract only or in combination with other natural contents
possess neuro-curative roles on different parts of the brain when exposed to different toxicants due to its rich
source of bioactive compounds (mainly polyphenols, nitric pigments, and saponins).11 While the health
benefits of carrot root are well-documented, the potential neuroprotective effects of its leaves remain largely
unexplored. However, preliminary research suggests that plant-derived antioxidants can effectively scavenge
free radicals, restore cellular homeostasis, and prevent neuronal damage,12 making them promising candidates
for mitigating heavy metal-induced neurotoxicity.13 This study aims to investigate the potential
neuroprotective role of carrot leaf extract (CLE) in cadmium-induced toxicity in the hippocampus and
prefrontal cortex of adult Wistar rats.
MATERIALS AND METHODS
A. Plant Collection and Identification Extract Preparation
Fresh carrot leaves were obtained from Ndufu Echara Community in Ikwo Local Government Area of Ebonyi
State, Nigeria. It was identified and authenticated by a Botanist in the Department of Science and
Biotechnology, University of Nigeria, Nsukka, with herbarium number 1017b.
Carrot leaves (2g) were collected and allowed to dry under shade for two weeks to prevent the direct effects of
sunlight on the active constituents of the leaves, after which they were grounded into powdery form in a
milling machine. The powder was sieved to obtain uniform particle size that was used in the extraction process
by the maceration method. The leaves were dissolved in ethanol at a ratio of 1:7, using 450 ml of ethanol. The
mixture was stirred every 6 hours over 48 hours. After this duration, it was sieved to extract the liquid content
and subsequently strained again using litmus paper. The obtained solution was dried with a water bath at 40°C,
and 2.5g, 5g and 7.5g of the extract were mixed with 30ml, 50ml and 70ml of water for the low dose, medium
dose and high dose treatment, respectively.14
B. Animal procurement
Thirty-five (35) Adult Wistar rats weighing 100-120g were purchased from the animal house of the study
institution. The animals were housed in well-ventilated wired cages and allowed to acclimatize for two weeks
in the animal house. They were maintained under standard photoperiodic conditions of 12 hours of light/dark
cycle at a temperature of 270C -300C and relative humidity of 50 ± 0.05C. The animals were fed with rat
pellets (Top Feed Ltd, Nigeria) and allowed unrestricted drinking water access.
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C. Experimental Design
Cadmium chloride was purchased from Zayo-Sigma Chemicals Ltd, Jos, Northern part of Nigeria, with
molecular weight 201.32g/mol, batch number 117 09, product number 80683 and pack size 250g, which was
used as the toxin for this experiment, the LD50 of cadmium is 88mg/kg. It was prepared using 1g of cadmium
which was dissolved in 50ml of distilled water. The constituted solution was shaken for proper dissolution and
then preserved in a refrigerator for use.
The study involved thirty-five (35) adult Wistar rats randomly divided into five (5) groups (Groups A-E), each
comprising seven (7) randomized rats, which were tagged and housed in separate cages. In preparing the stock
solution, 20g of CLE was dissolved in 100 ml of distilled water, from which subsequent concentrations for
administration were derived. According to Ijomone et al. (2020), 20 mg of Cadmium chloride (CdCl2) does not
cause morbidity.15 Furthermore, according to Ahmad et al. (2023), 800 mg/kg of CLE does not cause
morbidity.16
a. Group A (Control): Received normal rat feed and water only, orally for 28 days.
b. Group B received 8-mg/kg body weight (bwt) of CdCl2 orally for 28 days.
c. Group C was administered 400-mg/kg bwt of CLE orally for 28 days.
d. Group D received 8-mg/kg (bwt) of CdCl2 in saline and 200-mg/kg bwt of CLE for 28 days
e. Group E was administered 8-mg/kg bwt of CdCl2 in saline and 400-mg/kg bwt of CLE for 28 days.
D. Animal Sacrifice and Sample Collection
The animals were sacrificed at the end of 28 days using cervical dislocation after 24 hours of fasting, and
blood samples were collected from the apex of the heart for biochemical analysis while the skull was excised,
the prefrontal cortex and hippocampus were harvested and fixed in 10% formalin for histological studies.
E. Neurobehavioral Studies
Neuro-behavioural studies of the experimental animals were carried out using T-Maze, Sociability chamber
and Novel object recognition (NOR) test methods. These methods were used to ascertain the learning,
memory, and cognitive functions of the adult Wistar rats respectively.
F. Learning and Memory Test using T-Maze
The rats were subjected to a learning and memory test in a T-maze. The experiment was conducted according
to Maodaa et al. (2016).17 The rats were starved for 24h, given only water, before the experiment. The maze
was a wooden device consisting of three arms that formed a T shape: the right arm, the left arm, and the main
long arm. On the second day, animals were placed in the device for 3 minutes and allowed to explore each of
the arms. Afterwards, animals were returned to their cages for 3 hours and again placed in the T-maze for 5min
per animal.
Several parameters of behaviour, time spent in the left arm, number of entrances into the left arm, number of
entrances into the right arm, time spent in the right arm, number of entrances into the main arm, and time spent
in the main arm, were recorded for 5 min for each animal.
G. Novel Object Recognition Test
The NOR is specifically used to evaluate recognition memory and object recognition memory and is very
useful to study short-term memory, intermediate-term memory, and long-term memory via manipulation of the
retention interval.18 The task procedure consists of three phases; habituation, familiarization and test phase.
During the habituation phase, each Wistar rat was allowed to explore the open-field arena freely in the absence
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of objects and then removed from the arena and placed in its holding cage. During the familiarization phase, a
single rat was placed in the open-field arena containing two identical sample objects, of the same colour,
shape, texture and size, for a few minutes (retention interval) for the rats to familiarize themselves with the
objects. To prevent coercion to explore the objects, the rats were released against the centre of the opposite
wall with their backs to the objects. During the test phase, the rats were returned to the open-field arena with
two sample objects, one of which was familiar with, and the other the novel object (A + B).19 During the
familiarization and the test phase, the objects were placed in opposite and symmetrical corners of the open-
field arena and the location of novel versus familiar objects was counterbalanced.20 After the test phase, the
following parameters were collected namely; mean time spent sniffing familiar object, mean time spent
exploring familiar object, mean time spent exploring novel object, percentage of object discrimination (%), and
discrimination index
A. Calculation of discrimination index:
time with novel object − time with familiar object
time with novel object + time with familiar object
B. Calculation of Percentage of object discrimination (%):
(time with novel object)
time with novel object + time with familiar object
X 100
H. Immuno-histochemical Studies
Immuno-histochemical studies, was done using paraffin sections of the brain which were deparaffinised with
xylene, followed by antigen retrieval by heating in citrate buffer (10 mM, for 20 min). This was followed by
endogenous peroxidase blocking in 3% H2O2 for 10 min and incubated with rabbit anti-mouse rabbit anti-
GFAP (1:500 Santa Cruz Biotechnology).
After washing the slides with phosphate-buffered saline, the sections were incubated with suitable fluorescent
secondary antibodies, goat anti-rabbit AlexaFluor 488 (1:200) at room temperature for 1 hour, followed by
detection with 3-amino-9-ethylcarbazole, a chromogen. The slides were appropriately counterstained and
mounted in Paramount aqueous mounting medium.
Coronal sections were examined from the rostral anteroposterior (- 2.1 mm) to the anteroposterior (- 4.5 mm)
direction, as defined by the bregma of the brain atlas. Images were obtained at ×40 magnification using the
IMAGE PRO PLUS System (version 4.0; Media Cybernetics, Silver Spring, MD, USA) on a computer
attached to a light microscope (Zeiss Axioskop, Oberkochen, Germany), which interfaced with a charge-
coupled device video camera (Kodak Mega Plus model 1.4 I). Each image was stitched to avoid overlap of
adjacent tiles and exported in TIFF format
I. Immuno-labelling
Immuno-reactivity was quantified using the Image J software (National Institute of Health, USA). The
percentage of positively stained area in the hippocampus and cortex was measured for each section and 9 fields
were selected (randomly). The total field and immune-histochemical (IHC) stained areas were calculated and
the percentage of IHC stained area was calculated as follows:
Percentage of IHC stained area =
IHC stained area
Total area
X 100
J. Image Analysis of Immuno-Stained Slides
A digital bright-field microscope was used to examine immune-stained slides as photomicrographs were taken.
At x10 magnification, non-overlapping pictures of the hippocampus and prefrontal cortex were acquired and
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used for image analysis. Photomicrographs were analyzed and quantified using image analysis and processing
for Java (image J). The number of GFAP-positive cells were counted using the Image J cell counter.
K. Data Analysis
The experimental data obtained was analyzed using Graph Pad prism version 9.0.1.5 and results were
presented in a tabular form as Mean ± SEM. The significant differences between means were established using
a Two-Way Analysis of Variance (ANOVA). Multiple comparisons of Turkey’s Post Hoc Test were adopted
to check the significance level at p ≤ 0.05.
RESULTS
A. Body Weight Analysis
As shown in Table 1, group A, B, C, D and E had a weight changes of 10.30±3.94g (6.34±2.42%), -
22.90±4.11g (-12.84±2.31%), 9.40±1.98g (5.88±1.24%), -9.20±2.44g (-5.94±1.58%), and -0.80±2.61g (-
0.49±1.61%) respectively.
Table 1: Effects of Daucus carota ethanolic leaf extract on cadmium toxicity on animal weight change.
Groups Initial weight (g) Final weight (g) Weight Change (g) Percentage Weight Change (%)
A 162.50±5.96 172.80±9.90 10.30 ± 3.94 6.34 ± 2.42
B 178.40±5.98 155.50±10.09 -22.90 ± 4.11 -12.84 ± 2.31
C 159.90±3.66 169.30±4.51 9.40 ± 1.98 5.88 ± 1.24
D 154.90±4.25 145.70±6.69 -9.20 ± 2.44 -5.94 ± 1.58
E 162.00±3.93 161.20±6.54 -0.80 ± 2.61 -0.49 ± 1.61
No Significant difference.
B. Neuro-behavioural Study Analysis
The study showed the results of neurobehavioral studies of the animals assessed for Spatial memory using the
T-maze and recognition memory using the Novel object recognition tests, respectively as given below:
1). T-Maze Result
As shown in Table 2, in the T-maze test, TSIA A was 68.00±22.52 secs, 58.60±27.70 secs, 92.14±47.23 secs,
101.40±30.25 secs, and 99.36±25.29 secs in groups A to E respectively. TSIA B was 118.40±43.56 secs,
44.00±12.91 secs, 63.67±24.10 secs, 76.92±15.68 secs, and 76.92±15.68 secs in groups A to E respectively,
while TSNA was respectively 258.00±14.84 secs, 271.70±28.33 secs, 300.60±0.60 secs, 303.30±3.25 secs, and
303.30±3.25 secs in groups A to E. There was no significant difference among the groups.
Table 2: Effects of Daucus carota ethanolic leaf extract on cadmium neurotoxicity in rodent’s spatial memory.
Groups TSIA A (secs) TSIA B (secs) TSNA (sec)
A 68.00±22.52 118.40±43.56 258.00±14.84
B 58.60±27.70 44.00±12.91 271.70±28.33
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C 92.14±47.23 63.67±24.10 300.60±0.60
D 101.40±30.25 76.92±15.68 303.30±3.25
E 99.36±25.29 76.92±15.68 303.30±3.25
No Significant difference.
KEYS: TSIA A= Time spent in arm A; TSIA B= Time spent in arm B; TSNA= Time spent in no arm
2). Novel Object Recognition
In the novel object test, group B (cadmium only) showed a decrease in Tn (7.27±0.89s) and Tf (4.00±1.68s)
compared to group A (Control: Tn = 10.93±2.99s, Tf = 12.31±5.65s). Group C (200 mg/kg CLE) showed a
significant improvement (Tn = 21.65±3.23s, Tf = 23.96±3.04s, p<0.05) compared to group B. Groups D (Tn =
14.09±5.49s, Tf = 16.20±4.78s) and E (Tn = 7.37±2.52s, Tf = 7.62±3.13s) showed an increase in Tn and Tf
compared to group B, but the changes were not statistically significant as shown in table 3.
Table 3: Effects of Daucus carota ethanolic leaf extract in rodent’s impaired recognition memory due to
cadmium toxicity.
Groups Tn(secs) Tf(secs)
A 10.93±2.99 12.31±5.65
B 7.27±0.89 4.00±1.68
C 21.65±3.23 23.96±3.04b
D 14.09±5.49 16.20±4.78
E 7.37±2.52 7.62±3.13
a = Significant difference when compared to A; b = Significant difference when compared to B; c =
Significant difference when compared to C; d = Significant difference when compared to D; e = Significant
difference when compared to E.
KEYS: Tn - Time with Novel object; Tf = Time with Familiar object.
C. Effects of cadmium and carrot leaves extract on Neurodegenerative Proteins and apoptotic marker
As shown in Figures 1a and b, the Caspase-3 level was significantly increased in group B when compared to
the control group A. The carrot leave extract treated groups showed significant decrease in the level Caspase-3
when compared to group B at p<0.05 (Figure 1a). The results showed a significant decrease in the level of the
Tau proteins in group B compared to the control group A at p<0.05 (Figures 2b). The result also showed that
the extract treatment significantly increased the concentration of the Tau protein compared to the untreated
group B at p<0.05.
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Fig. 1: Effects of cadmium and carrot leaves extract on the levels of neurodegenerative protein and apoptotic
markers during the experiment. (a) Caspase-3 and (b) Tau protein. *Significant increase at p<0.05;
**Significant decrease at p<0.05; #Significant decrease at p<0.05; and ##Significant increase at p<0.05
D. Effects of cadmium and carrot leaves extract on the levels of Neurotransmitters
Regarding the histamine (Fig 2a), the levels were lowered significantly in group B when compared to the
control group A at p<0.05. The carrot leaves extract significantly increased the levels of histamine in the brain
when compared to group B at p<0.05. The level of Acetylcholine in the brain homogenate (Figure 2b), showed
that cadmium administration significantly increased the brain-acetylcholine concentration in group B when
compared to the control group A at p<0.05, while the level of acetylcholine was significantly lowered in all the
treated groups when compared to group B at p<0.05.
Fig 2: The effect of cadmium and carrot leaves extract on the levels of neurotransmitters (a) Histamine, and (b)
Acetylcholine. #Significant decrease at p<0.05 compared to A; ##Significant increase at p<0.05 compared to
B; *Significant increase at p<0.05 compared to A; **Significant decrease at p<0.05 compared to B
DISCUSSION
This study provided significant insights into the potential neuro-curative effects of Daucus carota ethanolic
leaf extract against cadmium-induced toxicity in the hippocampus and prefrontal cortex of adult Wistar rats.
The hippocampus and prefrontal cortex, which are essential for learning, memory, and executive functions are
particularly vulnerable to cadmium-induced damage.21,22
b
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A. Effects of CLE and Cd on the body weights
The body weight analysis highlights the detrimental effects of cadmium exposure and the potential curative
role of Daucus carota ethanolic leaf extract (CLE). As shown in table 1, the control group (group A) exhibited
normal weight gain (6.34%), consistent with expected physiological growth in Wistar rats. However, cadmium
exposure in group B resulted in significant weight loss (-12.84%), which aligns with a previous study that
reported reduced body weight in cadmium-exposed animals due to its toxic effects on metabolism and organ
function.23 Cadmium toxicity has been shown to disrupt lipid and protein metabolism, increase oxidative
stress, and impair nutrient absorption, all of which contribute to weight loss.24 In line with these findings,
Adegoke et al. (2017) also reported a significant decrease in final body weight following cadmium exposure.25
Contrary to this, another study has shown that low-dose cadmium exposure can lead to weight gain over time,
possibly due to metabolic compensatory mechanisms that increase adiposity following initial toxicity.26 These
variations may be influenced by the duration of exposure, dosage, and species differences.
Group C, which received CLE alone (400 mg/kg bwt), exhibited a weight gain of 5.88%, suggesting that CLE
does not induce weight loss under normal conditions but may instead support metabolic homeostasis. This
aligns with a previous study suggesting that carrot consumption has beneficial metabolic effects, with
carotenoids such as β-carotene helping to regulate adipose tissue function.27 Similarly, Mahesh et al. (2021)
reported that carrot juice is low in calories and rich in fiber, which may contribute to improved metabolic
function.28 Contradictory findings exist, as a study by Khan (2019) found no significant impact of carrot
extract on body weight.29 These discrepancies may be due to differences in the composition of the extract,
duration of administration, or the presence of other dietary factors influencing metabolism. Group D, which
was exposed to cadmium and treated with a lower dose of CLE (200 mg/kg bwt), showed weight loss (-
5.94%). Although this loss was less severe than in group B, it indicates that CLE at this dose only partially
mitigated cadmium-induced metabolic disruption. This aligns with previous study suggesting that while carrot
extracts possess antioxidant and anti-inflammatory properties, their ability to counteract cadmium toxicity may
be dose-dependent.30 However, it is important to note that some researches have implicated Daucus carota in
weight loss due to its fiber content, which promotes satiety and reduces caloric absorption.31 A study by
Ramirez et al. (2023) demonstrated that a high-fiber diet from carrot sources led to a reduction in body weight
by 10-12% over 12 weeks.30 This suggests that while CLE helps mitigate cadmium-induced weight loss, its
fiber content may also contribute to weight regulation by limiting energy intake.
Group E, which was exposed to cadmium and treated with a higher dose of CLE (400 mg/kg bwt), exhibited
minimal weight change (-0.49%), suggesting a stronger curative effect at this higher dose. This finding aligns
with a study indicating that plant extracts at higher concentrations may exert stronger anti-oxidative and
metabolic regulatory effects.32 The improved weight maintenance in this group may be attributed to the
digestion-stimulating properties of Daucus carota extract, which has been reported to enhance nutrient
absorption and counteract metabolic disturbances.32 Furthermore, a clinical study reported an inverse
relationship between fiber intake and BMI, fat oxidation, and energy storage, reinforcing the idea that Daucus
carota may support weight maintenance rather than promoting excessive weight gain.33
Comparing these findings with previous research highlights the variability in both cadmium’s and Daucus
carota's effects on body weight. While cadmium exposure in this study led to weight loss, some studies have
reported compensatory weight gain following prolonged exposure.26 Similarly, while CLE helped prevent
excessive weight loss in cadmium-exposed groups, its potential role in weight regulation remains complex.
Studies have reported both weight-maintaining and weight-reducing effects, likely due to variations in fiber
content, dosage, and metabolic response.30,32 The findings of this study suggest that while CLE provides some
ameliorating against cadmium-induced weight loss, its fiber content may also contribute to weight regulation,
depending on dosage and individual metabolic responses.
C. Neurobehavioral effects of CLE in Cadmium-induced toxicity
The neurobehavioral assessments in this study were conducted using the T-maze and novel object recognition
(NOR) tests to evaluate spatial and recognition memory, respectively. The results indicate that cadmium
exposure impaired recognition memory, while treatment with CLE demonstrated varying degrees of neuro-
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amelioration. In the T-maze test (table 2), TSIA A was slightly reduced in group B (cadmium-only) compared
to the control (group A), while groups C, D, and E exhibited prolonged times. The TSIA B was notably lower
in group B than in the control, indicating possible cadmium-induced deficits in spatial learning and decision-
making. Despite these differences, no statistically significant variations were found among the groups.
Previous research has established that cadmium exposure impairs spatial memory and learning ability in
rodents. Enogieru & Inegbedion (2022) reported that cadmium-treated rats displayed lower spontaneous
alternation behavior in maze tests, suggesting deficits in spatial working memory.34 Another study also
demonstrated that cadmium exposure leads to reduced exploratory behavior and impaired decision-making in
maze-based tests.35 In contrast, the lack of significant changes in the present study suggests that the cadmium
dose and duration of exposure may not have been sufficient to induce severe deficits in spatial memory.
Additionally, factors such as compensatory neuroplasticity or individual variability among animals may have
influenced the outcomes.
The NOR test (table 3) revealed a marked decrease in time spent with the novel object (Tn) and the familiar
object (Tf) in group B, indicating significant impairments in recognition memory due to cadmium exposure.
These findings are consistent with previous studies demonstrating that cadmium reduces exploratory behavior
and lowers discrimination index scores in rodents.35 The ability of cadmium to impair memory function has
been attributed to its neurotoxic effects, including oxidative stress, neuro-inflammation, and synaptic
dysfunction.36 Interestingly, group C (400 mg/kg CLE) showed a significant improvement in Tn and Tf,
suggesting that CLE at this dose provided notable neuro-amelioration against cadmium-induced memory
deficits. This aligns with findings that plant-derived antioxidants can mitigate neurotoxic damage and support
cognitive function. The improvement in group C may be attributed to the antioxidant properties of Daucus
carota, which has been shown to reduce oxidative stress and enhance synaptic plasticity.
Groups D (Cd + 200 mg/kg CLE) and E (Cd + 400 mg/kg CLE) showed moderate increases in Tn and Tf
compared to group B, but the changes were not statistically significant. While previous research on Daucus
carota and cognitive function is scarce, studies on other plant-based antioxidants have suggested that their
neuro-curative effects may be dose-dependent. It is possible that the higher CLE dose in groups D and E did
not provide additional cognitive benefits beyond those observed in group C. Alternatively, high concentrations
of certain bioactive compounds could have modulated neurotransmitter levels in a way that did not
significantly enhance recognition memory.
Cadmium has been shown to cross the blood-brain barrier, accumulate in the hippocampus, and cause neuronal
damage leading to cognitive impairment.37 Conversely, Akinyemi et al. (2017) demonstrated that cadmium
exposure caused severe recognition memory dysfunction, whereas Pulido et al. (2019) observed no statistical
differences in memory performance between cadmium-treated and control animals at certain time points.38,39
These discrepancies may be explained by differences in cadmium dosage, exposure duration, and species-
specific responses to heavy metal neurotoxicity.
Furthermore, behavioral alterations associated with cadmium exposure extend beyond memory deficits. A
study has shown that cadmium exposure can induce anxiety-like behaviors, social deficits, and altered
locomotor activity.40 Clinical evidence also suggests that cadmium exposure in humans is linked to learning
disabilities, hyperactivity, and lower IQ.41 The present findings contribute to this growing body of evidence by
demonstrating that cadmium impairs recognition memory, reinforcing its role as a neuro-toxicant with
cognitive consequences.
D. Effects of cadmium and carrot leaves extract on Neurodegenerative Proteins and apoptotic marker
This study (Fig 1a) showed that the level of Caspase-3 was significantly increased (at p<0.05) in group B
animals when compared to the control (group A). Meanwhile, the groups (C, D and E) treated with carrot
leave extract showed a significant decrease in the level of Caspase-3 when compared to group B at p<0.05.
This indicates the potentiality of carrot leave extract in ameliorating apoptosis. Therefore, carrot leave extract
exerted an inhibitory effect on the mitochondrial and death receptor pathways involved in Cd-induced
apoptosis. It has been suggested that Cd can cause apoptosis in a variety of cells, and can occur in a dose-and
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time-dependent manner.42 Pathak and Khandelwal (2006) reported that 6 h exposure to 25 μmol/l Cd induced
apoptosis in rat thymus cells.43 Caspase-3 is a frequently activated death protease, catalyzing the specific
cleavage of many key cellular proteins.44 Caspase-3 is a key factor in apoptosis, and is directly involved in the
chromosome condensation and DNA fragmentation processes.
Also, this study (Fig 1b) showed a significant decrease at p<0.05) in the level of the Tau proteins in group B
animals compared to the control (group A) while the treatment with carrot leave extract significantly increased
(at p<0.05), the concentration of the Tau protein compared to the untreated group B. This could mean that the
abnormal tau protein that accumulated as a result of cadmium exposure were not affected by administration of
carrot leaves extract. Meanwhile, the finding aligns with research suggesting that cadmium disrupts
cytoskeletal integrity by impairing tau protein phosphorylation.45 While CLE treatment in groups D and E
increased tau protein levels, these changes were not statistically significant, suggesting that further
investigation is needed to understand the precise role of Daucus carota in tau protein regulation.
Hyperphosphorylation of tau protein is proposed to be an early event for the evolution of tau pathology, and
may play an important role in Cd-induced neurodegeneration.46
E. Effects of cadmium and carrot leaves extract on the levels of Neurotransmitters
This study (Fig 2a) showed that the levels of histamine was lowered significantly (at p<0.05) in the cadmium
untreated animals (group B) when compared to the control (group A). However, the carrot leaves extract
significantly increased the levels of histamine in the brain when compared to group B at p<0.05. Recent
evidence suggests that aberrant histamine signaling in the brain may also be a key factor in addictive behaviors
and degenerative disease such as Parkinson's diseases and multiple sclerosis.47 Cadmium exposure has been
implicated to affect brain histamine levels and related signaling pathways, potentially contributing to neuro-
inflammation and neurotoxicity.48 Cadmium stimulates mast cell degranulation, leading to increased histamine
release and inflammatory mediator release, as well as disrupting redox signaling and calcium
influx.49 Furthermore, cadmium can alter the binding activity of histamine receptors in the brain, particularly
subtype 2, in certain regions.
Acetylcholine is a crucial neurotransmitter in the brain, playing a vital role in various cognitive functions like
memory, learning, and attention, as well as influencing mood and motivation.50 Cadmium exposure (Fig 2b)
significantly increased acetylcholine (ACH) levels in group B, indicating disrupted cholinergic
neurotransmission. Studies suggest that cadmium interferes with acetylcholine metabolism by inhibiting
acetylcholinesterase activity, leading to excessive ACH accumulation and neuronal hyperactivity.51 This
dysregulation has been implicated in cognitive impairments and neurodegenerative conditions.52 Meanwhile,
carrot leaf extract (CLE) treatment significantly reduced ACH levels in Groups C, D, and E, suggesting that it
may help restore cholinergic balance and improve cognitive function.
CONCLUSION
This study comprehensively evaluated the neurobehavioral and immune-histochemical effects of Daucus
carota ethanolic leaf extract in cadmium-induced toxicity of the hippocampus and prefrontal cortex of adult
wistar rats, assessing its impact on body weight, neurobehavior, oxidative stress markers, and
neurotransmitters in the hippocampus and prefrontal cortex. The findings confirmed that cadmium exposure
led to significant weight loss, cognitive impairments, increased oxidative stress, and severe neuronal
degeneration in a dose-dependent manner.
The observed neuronal regeneration in CLE-treated groups further support its neuro-ameliorating potential,
likely attributed to its rich antioxidant and anti-inflammatory bioactive compounds. While this study highlights
the potential of CLE as a natural therapeutic agent against cadmium-induced neurotoxicity, further research is
needed to explore its precise mechanisms of action and long-term effects in both experimental and clinical
settings.
Acknowledgment: We thank all our supervisors, staff of the Department of Anatomy in the study institution
for their valuable contributions to this research. We also appreciate our lecturers for their guidance and support
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ISSN No. 2321-2705 | DOI: 10.51244/IJRSI |Volume XII Issue VIII August 2025
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Ethical Approval
Ethical approval was sought and got from the Research Ethics Committee of the Faculty of Basic Medical
Sciences Ebonyi State University, Abakaliki, Ebonyi State, with number EBSU/REC/2024/7299. This research
also complied with the Helsinki declaration of 2013 as it concerns animal studies.
Conflict of Interests: The authors have no potential conflict of interests to declare
Data Availability Statement: All data about this research is available on reasonable request to the
corresponding author on ogashstanly90@yahoo.com
Author Contributions: The authors contributed equally to all aspects of this research
REFERENCES
1. Areba GO, Khalid R, Ngure RM, Maloba F, Nyaga N, Moseti KO, Ngotho M, Wanyoko JK, Karori
SM, Wachira FN. (2019). Neuroprotective effects of tea against cadmium toxicity. Bioactive
Compounds in Health and Disease, 2(12): 230–246. https://doi. org/10.31989/bchd.v2i12.684
2. Afzal A, Mahreen N. (2024). Emerging insights into the impacts of heavy metals exposure on health,
reproductive and productive performance of livestock. Frontiers in Pharmacology, 15:1375137.
https://doi.org/10.3389/fphar.2024.1375137
3. Rebello S, Sivaprasad MS, Anoopkumar AN, Jayakrishnan L, Aneesh EM, Narisetty V, Sindhu R,
Binod P, Pugazhendhi A, Pandey A.(2021). Cleaner technologies to combat heavy metal toxicity.
Journal of Environmental Management, 296:113231. https://doi.org/10.10 16/j.jenvman.2021.113231
4. Carmona A, Roudeau S, Ortega R.(2021). Molecular Mechanisms of Environmental Metal
Neurotoxicity: A Focus on the Interactions of Metals with Synapse Structure and Function. Toxics, 9(9):
198. https://doi.org/10.3390/toxics9090198
5. Du B, Zhou J, Lu B, Zhang C, Li D, Zhou J, Jiao S, Zhao K, Zhang H. (2020). Environmental and
human health risks from cadmium exposure near an active lead-zinc mine and a copper smelter, China.
Science of The Total Environment, 720: 137585. https://doi.org/10. 1016/j.scitotenv.2020.137585
6. Qing Y, Yang J, Zhu Y, Li Y, Zheng W, Wu M, He G. (2021). Dose-response evaluation of urinary
cadmium and kidney injury biomarkers in Chinese residents and dietary limit standards. Environmental
Health : A Global Access Science Source, 20(1):75. https://doi.org/ 10.1186/s12940-021-00760-9
7. Obeng-Gyasi E. (2019). Sources of lead exposure in various countries. Reviews on Environmental
Health, 34(1): 25–34. https://doi.org/10.1515/reveh-2018-0037
8. Siddiqua A, Hahladakis JN, Al-Attiya WAKA. (2022). An overview of the environmental pollution and
health effects associated with waste landfilling and open dumping. Environmental Science and
Pollution Research, 29(39): 58514–58536. https://doi.org/10.1007/s11 356-022-21578-z
9. Imafidon CE, Akomolafe RO, Eluwole OA, Adekunle IA, Agbaje RA. (2019). Aqueous garlic extract
improves renal clearance via vasodilatory/antioxidant mechanisms and mitigated proteinuria via
stabilization of glomerular filtration barrier. Clinical Phytoscience, 5(1):27.
https://doi.org/10.1186/s40816-019-0118-y
10. Ismail J, Shebaby WN, Daher J, Boulos JC, Taleb R, Daher CF, Mroueh M. (2023). The Wild Carrot
(Daucus carota): A Phytochemical and Pharmacological Review. Plants, 13(1): 93.
https://doi.org/10.3390/plants13010093
11. Matysek M, Kowalczuk-Vasilev E, Szalak R, Baranowska-Wójcik E, Arciszewski MB, Szwajgier D.
(2022). Can Bioactive Compounds in Beetroot/Carrot Juice Have a Neuroprotective Effect?
Morphological Studies of Neurons Immunoreactive to Calretinin of the Rat Hippocampus after
Exposure to Cadmium. Foods, 11(18): 2794. https://doi.org/10. 3390/foods11182794
12. Feng J, Zheng Y, Guo M, Ares I, Martínez M, Lopez-Torres B, Martínez-Larrañaga MR, Wang X,
Anadón A, Martínez MA. (2023). Oxidative stress, the blood-brain barrier and neurodegenerative
diseases: The critical beneficial role of dietary antioxidants. Acta Pharmaceutica Sinica. B, 13(10):
3988–4024. https://doi.org/10.1016/j.apsb. 2023.07.010
INTERNATIONAL JOURNAL OF RESEARCH AND SCIENTIFIC INNOVATION (IJRSI)
ISSN No. 2321-2705 | DOI: 10.51244/IJRSI |Volume XII Issue VIII August 2025
Page 2086
www.rsisinternational.org
13. Ru Q, Li Y, Chen L, Wu Y, Min J, Wang F. (2024). Iron homeostasis and ferroptosis in human diseases:
mechanisms and therapeutic prospects. Signal Transduction and Targeted Therapy, 9(1): 271.
https://doi.org/10.1038/s41392-024-01969-z
14. Owolabi OJ, Amaechina FC, Okoro, M. (2011). Effect of Ethanol Leaf Extract of Newboulda Laevis on
Blood Glucose Levels of Diabetic Rats. Tropical Journal of Pharmaceutical Research, 10(3): 249–254.
https://doi.org/10.4314/tjpr.v10i3.12
15. Ijomone OM, Ifenatuoha CW, Aluko OM, Ijomone OK, Aschner M. (2020). The aging brain: Impact of
heavy metal neurotoxicity. Critical Reviews in Toxicology, 50(9): 801–814.
https://doi.org/10.1080/10408444.2020.1838441
16. Ahmad T, Cawood M, Iqbal Q, Ariño A, Batool A, Tariq RMS, Azam M, Akhtar S.(2019).
Phytochemicals in Daucus carota and Their Health Benefits-Review Article. Foods (Basel,
Switzerland), 8(9): 424. https://doi.org/10.3390/foods8090424
17. Maodaa SN, Allam AA, Ajarem J, Abdel-Maksoud MA, Al-Basher GI, Wang ZY. (2016). Effect of
parsley (Petroselinum crispum, Apiaceae) juice against cadmium neurotoxicity in albino mice (Mus
Musculus). Behavioral and Brain Functions, 12(1): 6. https://doi.org/10. 1186/s12993-016-0090-3
18. Taglialatela G, Hogan D, Zhang WR, Dineley KT. (2009). Intermediate- and long-term recognition
memory deficits in Tg2576 mice are reversed with acute calcineurin inhibition. Behavioural Brain
Research, 200(1): 95–99. https://doi.org/10.1016/j.bbr.2008.12. 034
19. Gaskin S, Tardif M, Cole E, Piterkin P, Kayello L, Mumby DG. (2010). Object familiarization and
novel-object preference in rats. Behavioural Processes, 83(1): 61–71. https://doi.org/
10.1016/j.beproc.2009.10.003
20. Hammond RS, Tull LE, Stackman RW. (2004). On the delay-dependent involvement of the
hippocampus in object recognition memory. Neurobiology of Learning and Memory, 82(1): 26–34.
https://doi.org/10.1016/j.nlm.2004.03.005
21. Friedman NP, Robbins TW. (2022). The role of prefrontal cortex in cognitive control and executive
function. Neuropsychopharmacology, 47(1), 72–89. https://doi.org/10.1038/s 41386-021-01132-0
22. Yu G, Wu L, Su Q, Ji X, Zhou J, Wu S, Tang Y, Li, H.(2024). Neurotoxic effects of heavy metal
pollutants in the environment: Focusing on epigenetic mechanisms. Environmental Pollution, 345:
123563. https://doi.org/10.1016/j.envpol.2024.123563
23. Padilla MA, Elobeid M, Ruden DM, Allison DB. (2010). An examination of the association of selected
toxic metals with total and central obesity indices: NHANES 99-02. International Journal of
Environmental Research and Public Health, 7(9): 3332–3347. https://doi.org/10.3390/ijerph7093332
24. Nazimabashir, Manoharan, V, Prabu SM. (2014). Protective role of grape seed proanthocyanidins
against cadmium induced hepatic dysfunction in rats. Toxicology Research, 3(2), 131.
https://doi.org/10.1039/c3tx50085c
25. Adegoke A, Salami A, Olaleye S. (2017). Cadmium Exacerbates Acetic Acid Induced Experimental
Colitis in Rats. European Journal of Experimental Biology, 7(5): 27. https://doi.org/10.21767/2248-
9215.100027
26. Sarmiento-Ortega VE, Alcántara-Jara DI, Moroni-González D, Diaz A, Vázquez-Roque RA, Brambila
E, Treviño S. (2025). Chronic cadmium exposure to minimal-risk doses causes dysfunction of
epididymal adipose tissue and metabolic disorders. Toxicology and Applied Pharmacology, 495:
117203. https://doi.org/10.1016/j.taap.2024.117203
27. Fujihara K, Nogawa S, Saito K, Horikawa C, Takeda Y, Cho K, Ishiguro H, Kodama S, Nakagawa Y,
Matsuzaka T, Shimano H, Sone H. (2021). Carrot Consumption Frequency Associated with Reduced
BMI and Obesity through the SNP Intermediary rs4445711. Nutrients, 13(10): 3478.
https://doi.org/10.3390/nu13103478
28. Mahesh M, Pandey H, Raja Gopal Reddy M, Prabhakaran Sobhana P, Korrapati D, Uday Kumar P,
Vajreswari A, Jeyakumar SM. (2021). Carrot Juice Consumption Reduces High Fructose-Induced
Adiposity in Rats and Body Weight and BMI in Type 2 Diabetic Subjects. Nutrition and Metabolic
Insights, 14: 11786388211014916. https://doi.org/10.1177/ 11786388211014917
29. Khan SR. (2019). Utilization Carrot Pulp as Corn Replacement in the Broiler Diet. IOSR Journal of
Agriculture and Veterinary Science (IOSR-JAVS), 12(2): 72–74. https://doi.org/ 10.9790/2380-
1202027274
INTERNATIONAL JOURNAL OF RESEARCH AND SCIENTIFIC INNOVATION (IJRSI)
ISSN No. 2321-2705 | DOI: 10.51244/IJRSI |Volume XII Issue VIII August 2025
Page 2087
www.rsisinternational.org
30. Ramirez MR, Manuale D, Yori JC. (2023). Assessment of effectiveness of oral supplementation of
isolated fiber of carrot on metabolic parameters in mature rats. Food Science and Human Wellness,
12(6): 2022–2028. https://doi.org/10.1016/j.fshw.2023.03. 016
31. El-Anany AM, Ali RFM. (2018). Hypolipidemic effect of coffee silver skin in rats fed a high-fat diet.
Food Science and Human Wellness, 7(4): 252–259. https://doi.org/10.1016 /j.fshw.2018.10.005
32. Dabai SA, Bello S, Dabai JS. (2023). Growth performance and carcass characteristics of finisher
broiler chickens served carrot leaf extract as a supplementary source of vitamins and minerals. Nigerian
Journal of Animal Science, 23(1), 144–149.
33. He K, Hu FB, Colditz GA, Manson JE, Willett WC, Liu S. (2004). Changes in intake of fruits and
vegetables in relation to risk of obesity and weight gain among middle-aged women. International
Journal of Obesity, 28(12): 1569–1574. https://doi.org/10.1038 /sj.ijo.0802795
34. Enogieru AB, Inegbedion GO. (2022). Attenuation of Oxidative Stress and Cognitive Impairment in
Cadmium Chloride-Exposed Wistar Rats Pre-treated with Ethanolic Turmeric Root Extract. The
Journal of Phytopharmacology, 11(2): 118–124. https://doi.org/10.312 54/phyto.2022.11212
35. Kim K, Melough MM, Vance TM, Noh H, Koo SI, Chun OK. (2018). Dietary Cadmium Intake and
Sources in the US. Nutrients, 11(1): 2. https://doi.org/10.3390/nu11010002
36. Mahdavi S, Khodarahmi P, Roodbari NH. (2018). Effects of cadmium on Bcl-2/ Bax expression ratio in
rat cortex brain and hippocampus. Human & Experimental Toxicology, 37(3): 321–328.
https://doi.org/10.1177/0960327117703687
37. Favorito R, Monaco A, Grimaldi MC, Ferrandino I. (2017). Effects of cadmium on the glial
architecture in lizard brain. European Journal of Histochemistry : EJH, 61(1): 2734.
https://doi.org/10.4081/ejh.2017.2734
38. Akinyemi AJ, Oboh G, Fadaka AO, Olatunji BP, Akomolafe S. (2017). Curcumin administration
suppress acetylcholinesterase gene expression in cadmium treated rats. Neurotoxicology, 62: 5–79.
https://doi.org/10.1016/j.neuro.2017.05.004
39. Pulido G, Treviño S, Brambila E, Vazquez-Roque R, Moreno-Rodriguez A, Peña Rosas U, Moran-
Perales JL, Handal Silva A, Guevara J, Flores G, Diaz A. (2019). The Administration of Cadmium for
2, 3 and 4 Months Causes a Loss of Recognition Memory, Promotes Neuronal Hypotrophy and
Apoptosis in the Hippocampus of Rats. Neurochemical Research, 44(2): 485–497.
https://doi.org/10.1007/s11064-018-02703-2
40. Alanazi MM, Ansari MA, Nadeem A, Attia SM, Bakheet SA, Al-Mazroua HA, Aldossari AA,
Almutairi MM, Albekairi TH, Hussein MH, Al-Hamamah MA, Ahmad SF. (2023). Cadmium Exposure
Is Associated with Behavioral Deficits and Neuroimmune Dysfunction in BTBR T(+) Itpr3tf/J Mice.
International Journal of Molecular Sciences, 24(7): 6575. https://doi.org/10.3390/ijms24076575
41. Xu C, Chen S, Xu M, Chen X, Wang X, Zhang H, Dong X, Zhang R, Chen X, Gao W, Huang S, Chen
L. (2021). Cadmium Impairs Autophagy Leading to Apoptosis by Ca2+-Dependent Activation of JNK
Signaling Pathway in Neuronal Cells. Neurochemical Research, 46(8): 2033–2045.
https://doi.org/10.1007/s11064-021-03341-x
42. Dong S, Shen HM and Ong CN. (2001). Cadmium-induced apoptosis and phenotypic changes in
mouse thymocytes. Mol Cell Biochem 222: 11-20.
43. Pathak N and Khandelwal S. (2006). Modulation of cadmium induced alterations in murine thymocytes
by piperine: Oxidative stress, apoptosis, phenotyping and blastogenesis. Biochem Pharmacol 72:
486-497.
44. Porter AG, Jänicke RU. (1999). Emerging roles of caspase-3 in apoptosis. Cell Death Differ. 6(2):99-
104.
45. Wang B, Du Y. (2013). Cadmium and its neurotoxic effects. Oxidative Medicine and Cellular
Longevity, 2013: 898034. https://doi.org/10.1155/2013/898034
46. Peiling B, Zhengping Z, Yanyan Z, Aiying X, Yanhong G, Jianyun M, Zhimin Y, Lan Ll (2016).
Theanine attenuates cadmium-induced neurotoxicity through the inhibition of oxidative damage and tau
hyperphosphorylation, NeuroToxicology, 57: 95-103.
47. Passani MB, Panula P, Lin JS. (2014). Histamine in the brain. Front Syst Neurosci. 28(8):64.
48. Giusi G, Facciolo RM, Alò R, Carelli A, Madeo M, Brandmayr P, Canonaco M. (2005). Some
environmental contaminants influence motor and feeding behaviors in the ornate wrasse (Thalassoma
pavo) via distinct cerebral histamine receptor subtypes. Environ Health Perspect.113(11):1522-9.
INTERNATIONAL JOURNAL OF RESEARCH AND SCIENTIFIC INNOVATION (IJRSI)
ISSN No. 2321-2705 | DOI: 10.51244/IJRSI |Volume XII Issue VIII August 2025
Page 2088
www.rsisinternational.org
49. Vervloet D, Okazaki T, Llea VS, Reisman RE, Arbesman CE. (1975). Histamine release from human
leukocytes: modulation by cadmium ion. J Allergy Clin Immunol. 56(4):262-72.
50. Mineur YS, Picciotto MR. (2023). How can I measure brain acetylcholine levels in vivo? Advantages
and caveats of commonly used approaches. J Neurochem. 167(1):3-15.
51. Del Pino J, Zeballos G, Anadon MJ, Capo MA, Díaz MJ, García J, Frejo MT. (2014). Higher sensitivity
to cadmium induced cell death of basal forebrain cholinergic neurons: a cholinesterase dependent
mechanism. Toxicology, 325: 151–159. https://doi.org/10. 1016/j.tox.2014.09.004
52. Lamtai M, Azirar S, Zghari O, Ouakki S, El Hessni A, Mesfioui A, Ouichou A. (2021). Melatonin
Ameliorates Cadmium-Induced Affective and Cognitive Impairments and Hippocampal Oxidative
Stress in Rat. Biological Trace Element Research, 199(4): 1445–1455. https://doi.org/10.1007/s12011-
020-02247-z