This study aimed to investigate the neuroprotective effects of Synclisia scabrida on colorant induced neurodegeneration in male wistar rats. The specific objectives include; • To determine the hippocampal and prefrontal cortex antioxidant activities (SOD, MDA, Catalase). The rate at which food colorants are consumed in Nigeria today is on the increase. Studies have shown that excessive intake of food colorants has detrimental effects on the brain and other vital organs like the kidney (Wopara, et al., 2021). Hence, this study was to ascertain the ameliorative effects of methanolic extract of S. scabrida on neurodegeneration caused by the colorants (tartrazine and erythrosine). Traditional medicine including the use of natural products from plants is widely practiced around the world. Certain natural products contain enhancing and healing properties that may in fact have an essential role in the human system. The parameters assessed in this study include the hippocampal and prefrontal cortex SOD, MDA, CAT and the histology of the hippocampus and prefrontal cortex. Antioxidants are important in promoting the well-being of humans. They are important in the promotion of human health by neutralizing cellular damage caused by free radicals or reactive oxygen species. The formation of reactive oxygen species is as a result of most varieties of essential biological reactions. It is also known that oxygen radicals could be formed in excess in most chronic disease (Elzbieta, et al., 2005). The consumption of food colorants caused neurodegeneration by altering the brain antioxidant markers. The administration of methanolic leave extract of Synclisia scabrida led to the increased brain antioxidant concentrations (SOD, MDA, CAT), reduced neurodegeneration, increased neuroprotection and decreased lipid peroxidation. These effects occurred particularly in group fed with high dose of extract. It is of the view that the neurodegeneration seen in this study especially in group 2 may likely be due to oxidative stress. This could be linked to increased brain antioxidant enzymes coupled with the production of huge quantities of free radicals relative to the little quantity of antioxidant enzymes present to get rid of them. However, consideration and precaution are advised in the intake of food colorants and additives as sweeteners. We contribute that intake of S. scabrida increased the brain antioxidant concentration and could ameliorate the harmful effects of food colorants.

1. EFFECTS OF Synclisia scabrida ON NEUROPROTECTION: AN AMELIORATIVE
STUDY ON COLORANT INDUCED NEURODEGENERATIVE TOXICITY ON MALE
WISTAR RATS
BY
EMERUEM AUGUSTINE CHIDERA
PHY/T/18/1745
DEPARTMENT OF HUMAN PHYSIOLOGY
FACULTY OF BASIC MEDICAL SCIENCES,
MADONNA UNIVERSITY ELELE CAMPUS RIVERS STATE, NIGERIA.
SUPERVISOR: DR. MOBISSON S.K.

2. BACKGROUND OF STUDY
 It has been documented that some food additives like erythrosine and tartrazine
causes neurodegeneration by negatively influencing the brain cells (Wopara, et
al., 2021). Some studies have shown that Synclisia scabrida have
neuroprotective potentials which has been attributed to its phytonutrients
(Samuel, 2018).
 Synclisia scabrida (hot leaf) is a wild shrub belonging to the family of
Menispermaceae which is commonly found in different parts of Africa where it
occurs chiefly in rainforests at medium and low altitude (Nnamudi, et., al
2020).
Synclisia scabrida has ethnomedicinal uses in many parts of Africa, especially
Nigeria, Cameroon, Gabon and Congo. The root, leaf, stem and liana are used
to treat gastric ulcer, malaria, mental disorders, venereal diseases, prostate
problems, asthma, hernia, upper abdominal pains, dyspepsia, menstrual pains,
prevent spontaneous abortion and as an aphrodisiac (Wang, et., al 2018).
Pictorial representation of Synclisia
scabrida (Hot leaf)

3. BACKGROUND OF STUDY
 Erythrosine is an artificial red (cherry-pink) food coloring made from coal
tar. It is an organic compound containing iodine and
sodium. Erythrosine is also commonly referred to as red dye.
The red food dye erythrosine contains high levels of bound iodine,
although there is evidence that the bioavailability of this iodine is relatively
low (Wenlock, et al., 1982).
The harmful chemical is found to be detrimental to brain health as well.
Experts have warned that erythrosine has a negative effect on yeast cells
and is known to cause phototoxicity or hypersensitivity to light.

4. BACKGROUND OF STUDY
Tartrazine, also referred to as FD&C yellow #5, is an artificial
(synthetic) food dye. It is one of several azo food dyes that are
made from petroleum products.
Tartrazine appears to be a neurotoxin (toxic to cells in the brain) at
least in rats (Fangxu, et al., 2010).

5. BACKGROUND OF STUDY
 The nervous system is a highly complex part of an animal that coordinates
its action and sensory information by transmitting signals to and from
different part of its body (Tortora, et al., 2016).
 The nervous system detects environmental changes that impact the body,
then works in tandem with the endocrine system to respond to such events.
In vertebrates it consists of two main parts, the central nervous
system (CNS) and the peripheral nervous system (PNS).

6. STATEMENT OF PROBLEM
The increased rate at which food colorant is consumed has shown great
effect on the neurodegeneration. This work is aimed at investigating the
neuroprotective effects of Synclisia scabrida.

7. JUSTIFICATION OF STUDY
The increasing population of people suffering from neuronal
disorders in Nigeria is increasing by the day, and it has shown that
natural plants like Synclisia scabrida may possess potential remedies
for neurodegeneration.

8. AIM OF STUDY
This study was aimed to investigate the neuroprotective effects of
Synclisia scabrida on colorant induced neurodegeneration in male
wistar rats.

9. STUDY OBJECTIVES
The objectives of this study include;
 To determine the hippocampal and prefrontal cortex antioxidant
activities (SOD, MDA, Catalase).

10. MATERIALS AND METHODS
Materials, Reagents and Equipment
 The materials that were used for this study are as follows;
 Aluminum cages and saw dust, water cans, feeding troughs, syringes, micro capillary
tube, orogastric cannula, plain sample bottles, EDTA sample bottles, test tubes of
various sizes, normal saline, dissecting set, dissecting board, water bath, distilled
water, cotton wool, medicated soap, chloroform anesthesia, desiccator bottle, slides
and cover slips, manual animal weighing balance and electronic weighing (weighs up
to 0.001).

11. MATERIALS AND METHODS
STUDY ANIMALS
 A total of twenty (20) male wistar rats, aged 5 weeks and weighing 190- 210g were
used for this study.
The animals were purchased and housed in the department of physiology, Madonna
University. Animal House and were allowed to acclimatized for 2 weeks and exposed to
12/12 hours light/dark cycle.
Animals had access to feed and water ad libitum. The animals were kept in line with laid
down principles for animal care as prescribed in Helsinki`s 1964 declaration.

12. MATERIALS AND METHODS
Treatments:
The animals, after 2 weeks of acclimatization were housed In clean
cages in a room temperature of 23oC to 25oC.
They were fed with standard feed and water.
No animal was subject to pain, suffering, or restraint.
They were maintained in accordance to the guidelines set for the care
of laboratory animals. (DHIS, NIH publication No. 85-23, 1985).

13. MATERIALS AND METHODS
Administration
 Administration of combined dose of the colorants and S. scabrida,
were done via oral route, with the help of an orogastric tube (gavage),
to animals in the treatment groups (2-4). The administration was for
30days, and after which the animals were sacrificed, blood sample
collected and the whole brain was harvested and fixed for 12hrs with
boiun’s saline. Thereafter, the hippocampus and prefrontal cortex was
isolated for brain oxidative stress marker assay.

14. Groups No. of rats Treatment
Group A
(Control)
5 Received feed and 0.5 ml of Normal saline as a vehicle throughout the
experiment.
Group B
(Negative
control)
5 Received 20mg/kg body weight of combined dose of colorants
(erythrosine and tartrazine), no extract was given.
Group C 5 Received 20mg/kg body weight of colorant + 25mg/kg body weight of
extract.
Group D 5 Received 20mg/kg body weight of colorant + 50mg/kg of extract.

15. MATERIALS AND METHODS
SAMPLE COLLECTION:
The animals were anaesthetized with Diethyl-ether.
 Brain samples were collected for biochemical assay and histology, the
histological brain sample was fixed in 10% formal saline, whereas, the brain
sample for biochemical assay was fixed in 5ml normal saline.

16. MATERIALS AND METHODS
Brain Tissue Homogenization:
 All animals were anaesthetized with diethyl-ether and perfused with saline
(0.9% NaCl) followed by 4% paraformaldehyde (PFA) in phosphate buffer
(PB;0.1 M; pH 7.4)
All extracranial tissues were removed, the hippocampus’s and prefrontal
cortex’s were assay from the brain
After overnight post fixation of 4oC, the hippocampus and prefrontal cortex’s
assayed from the brains were stored in formaldehyde and placed in a
refrigerator (4oC) until it was used for biochemical analysis.

17. MATERIALS AND METHODS
Statistical analysis:
 The multiple comparisons were done using one way
ANOVA and POST HOC test using LSD. The values
were presented as Mean±SEM and(P<0.05) was
considered statistically significant.

18. RESULTS
0.022
0.027
0.038
[0.059] *
0
0.01
0.02
0.03
0.04
0.05
0.06
0.07
GRP 1 GRP 2 GRP 3 GRP 4
SOD
Hippocampus
Groups
SOD HPC
FIG.1 Hippocampal SOD Concentration in different experimental groups
Values presented as mean± SEM. P-value =*P<0.05 VS Control, N=5

19. 0.119
0.144*
0.135
0.151
0
0.02
0.04
0.06
0.08
0.1
0.12
0.14
0.16
0.18
GRP 1 GRP 2 GRP 3 GRP 4
MDA
Hippocampus
Groups
MDA HPC
FIG.2 Hippocampal MDA Concentration in different experimental groups
Values presented as mean± SEM. P-value =*P<0.05 VS Control, N=5
RESULTS

20. 2.64
1.364*
3.878*
4.468*
0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
5
GRP 1 GRP 2 GRP 3 GRP 4
CAT
Hippocampus
Groups
CATHPC
FIG.3 Hippocampal CAT Concentration in different experimental groups
Values presented as mean± SEM. P-value =*P<0.05 VS Control, N=5
RESULTS

21. 0.06 0.059
0.071
0.067
0
0.01
0.02
0.03
0.04
0.05
0.06
0.07
0.08
1 2 3 4
SOD
Prefrontal
cortex
Groups
SOD PFC
FIG.4 Prefrontal Cortex SOD Concentration in different experimental groups
Values presented as mean± SEM. P-value =*P<0.05 VS Control, N=5
RESULTS

22. 0.036
0.06
0.072*
0.083*
0
0.01
0.02
0.03
0.04
0.05
0.06
0.07
0.08
0.09
GRP 1 GRP 2 GRP 3 GRP 4
MDA
Prefrontal
cortex
Groups
MDA PFC
FIG.5 Prefrontal Cortex MDA Concentration in different experimental groups
Values presented as mean± SEM. P-value =*P<0.05 VS Control, N=5
RESULTS

23. 1.154
0.394*
1.44
2.648*
0
0.5
1
1.5
2
2.5
3
GRP 1 GRP 2 GRP 3 GRP 4
CAT
Prefrontal
cortex
Groups
CAT PFC
FIG. 6 Prefrontal Cortex CAT Concentration in different experimental groups
Values presented as mean± SEM. P-value =*P<0.05 VS Control, N=5
RESULTS

24. SUMMARY OF FINDINGS
In this study, the concentration of the hippocampal and prefrontal cortex antioxidants
enzymes (SOD, MDA, CAT) was significantly increased (p<0.05) in Synclisia scabrida
extract fed rats. The significant increase in antioxidant activity may be attributed to
increase oxidative stress caused by the colorants (Wopara, et al., 2021).
This study was to ascertain the ameliorative effects of methanolic extract of S. scabrida
on neurodegeneration caused by the colorants (tartrazine and erythrosine). The
parameters assessed in this study include the hippocampal and prefrontal cortex SOD,
MDA, CAT and the histology of the hippocampus and prefrontal cortex.

25. SUMMARY OF FINDINGS
The increase in the concentration of these brain antioxidative enzymes
within the hippocampus and the prefrontal cortex of colorants fed rats
(erythrosine and tartrazine) indicates a high level of lipid peroxidation
which would have great impacts in the damage of cell membrane layer,
this would eventually lead to the damage of the brain cells causing
neurodegeneration.
Hence, the increase in brain antioxidant concentration in this study
may likely be an indication of reduced oxidative stress generated by
reactive oxygen species and to decrease neurodegeneration.

26. CONCLUSION
The consumption of food colorants caused neurodegeneration by altering the brain
antioxidant markers. The administration of methanolic leave extract of Synclisia scabrida
led to the increased brain antioxidant concentrations (SOD, MDA, CAT), reduced
neurodegeneration, increased neuroprotection and decreased lipid peroxidation.
These effects occurred particularly in group fed with high dose of extract. It is of the view
that the neurodegeneration seen in this study especially in group 2 may likely be due to
oxidative stress. Hence, consideration and precaution should be taken in the intake of food
colorants and additives as sweeteners.

27. CONTRIBUTION TO KNOWLEDGE
A. The consumption of food colorants (tartrazine and erythrosine) causes
neurodegeneration in rats.
B. Consumption of methanolic extract of S. scabrida caused an increase in
the concentration of SOD, CAT and MDA in rats.
C. Ingestion of methanolic extract of S. scabrida may have neuroprotective
effects on the rats.

28. RECOMMENDATION
This work is animal-based study, it is recommended that similar study is
replicated in humans.

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30. Thank You