INTRODUCTION
Clove
(Syzygium aromaticum)
is one of the most commonly used spices around the world including Nigeria in
Africa as preservative, medicine as well as in the preparation of various spicy
rich dishes. The clove tree is a perennial tropical plant which grows to a
height of about 10- 20meters, having large oval leaves and crimson flowers in
numerous groups of terminal clusters. Clove bud is the dried, unopened,
nail-shaped, flower bud of the evergreen Syzygium aromaticum tree. Clove bud is aromatic,
a stimulant and carminative and is used for dyspepsia and gastric irritation
(Ahmed et al, 1986). In addition to its flavoring, pungent and fragrant
properties as well as culinary uses, the clove bud and its oil have an
abundance of medicinal and recreational uses. The use of clove bud in folk
medicine as diuretic, odontalgic, stomachic, tonicardiac, aromatic condiment and condiment with
carminative and stimulant activity has been reported (Boulos
1983). It is also used in dentistry as a natural analgaesic
and antiseptic due to its possession of eugenol; its
major component (Rathi et al., 2012). Clove oil; an
essential oil derived from the dried flower buds, leaves and stems of Syzygium aromaticum
tree has a spicy aroma which makes it a sought after ingredient for food
flavoring, fragrance formulations as well as in the production of soaps and
detergents. The insecticidal properties of clove oil have been studied against
lice, weevils, moth caterpillars, beetles and cockroaches (El-Raouf et al., 2019). It has also been found to possess
mosquito-repelling properties (Saurabh et al., 2024)
and (Bahmanpour et al., 2015) Clove oil contains
several compounds with eugenol been the major
component. Other compounds that have been determined to be present in clove oil
include β-caryophyllene, eugenyl
acetate, α-humulene and others. The oil of clove
has also been reported to be a good remedy for bronchitis, the common 2 cold
cough, fever and sore throat. Clove buds extracts and especially its oil have
been reported to possess anti-oxidant, anti-fungi, antiviral, anti-microbial,
anti-diabetic, anti-inflammatory, anti-thrombotic, anesthetic, pain-relieving
and insect repellent properties (Pramod et al., 2010)
and (Vaney et al., 2024) Some Polyphenols such as
Gallic acid, Ellagic acid, Quercetin
glycoside, Ellagic acid derivative and some
unidentified phenolic compounds have been characterized and reported to be
present in clove buds( Ahmed et al., 2016) The reproductive health is a state
of complete physical, mental and social well-being and not merely the absence
of disease or infirmity, all matters relating to the reproductive system and
its function and processes. Reproductive health implies that people are able to
have a satisfying and safe sex and they have the capability to reproduce and
the freedom to decide if, and how often to do so. (According to WHO). In women
the reproductive health depends on maintaining coordinated responses of network
of endocrine signals that functions primarily to ensure successful procreation
but also have other wide- ranging influences on the female body, including not
only secondary reproductive tissues such as the breast, but also nonreproductive tissues such as the bone, brain and
cardiovascular system. Reproductive health is a crucial aspect of overall
wellbeing, and any disruption or imbalance in the reproductive system can have
significant implications for fertility and overall health in females.
Understanding the effects of natural substances, such as cloves, on the
reproductive parameters is therefore important for both clinical and research
purposes. Previous studies have demonstrated the estrogenic properties of
certain phytochemicals found in cloves, such as eugenol
and flavonoids. Estrogen is a key hormone in the regulation of female reproductive
functions and is involved in a wide range of processes, including the growth
and development or reproductive organs, follliculogenesis,
ovulation, and maintenance of pregnancy. The intricate relationship between
natural substance and reproductive health has 3 been a subject of growing
interest in scientific inquiry. Understanding the effect of graded doses of
cloves on the uterus and fallopian tubes is of particular importance in
elucidating potential implications for reproductive health. The uterus, a central
organ in female reproductive system plays a pivotal role in fertility and
gestation, while the fallopian tube facilitates the crucial early stage of
embryonic development investigating the impact of cloves on these vital
components offers valuable insight into then influence on female reproductive
physiology.
MATERIALS AND METHODS
Animal care
A
total of twenty healthy wistar rats weighing
150-200grams were used in the study and they were acquired from laboratory
animal house of University of Port Harcourt. The animals were housed in a
hygienic and well ventilated compartment maintained under standard
environmental conditions and fed with normal rodent chew and allowed access to
drinking water for fourteen days to allow acclimatization before the
commencement of the experiment.
Preparation of
extract
A
total five grams (5gm) of the powdered herbs and 20mL of distilled water were
added into conical flasks. The flasks were covered with a wooden cork and
contents of the flasks were mixed thoroughly. The flasks were placed at shaker
adjusted at 100 rpm overnight. The mixtures were then filtered through a muslin
cloth and centrifuged at 2000rpm for 5 min and supernatant was transferred into
sterile falcon tube after filtration and stored at 4°C in refrigerator.
Ethical Approval
Ethical
consideration was gotten from the Board of The Department of Human Physiology Bayelsa Medical University, Yenagoa,
Bayelsa State.
Experimental
procedure:
After
two weeks of acclimatization, the rats were weighed and assigned to their
respective groups. The twenty-adult female wistar
rats were allocated into four groups (1, 2, 3 and 4) of five animals each.
During the experiment, Group 1 served as the control group receiving no extract
while Groups 2, 3 and 4 each received 1ml, 1.5ml and 2ml each day respectively
for a period of 28 days by oral administration and the weight of the animals
were recorded twice a week. During the treatment period, the rats were closely
monitored for any signs of distress or adverse effects. Group 1 (Control group)
receiving no cloves just distilled water. Group 2, received low dose of cloves
extract. Group 3, received moderate dose of clove extract while Group 4,
received high dose of clove extract.
Collection of samples
At
the end of the experimental period, the animals were allowed to fast for 12
hours and thereafter they were sacrificed. The organs from the rats were
harvested and blood samples were collected by cardiac puncture using sterile
needles and syringes. Portion of blood were transferred in a sterile plain
sample bottles and used for the determination of oxidative parameters and
hormonal analysis. Blood samples will be analyzed for levels of estrogen,
progesterone using ELISA or similar techniques. And for the oxidative stress,
tissue samples will be used to measure oxidative stress markers such as
superoxide dismutase, malondialdehyde levels as a
lipid peroxidation marker.
Statistical Analyses
Version
21.0 of the IBM Statistical Product and Service Solutions (SPSS) software was
used to analysis the quantitative data obtained from the present study. One-way
analysis of variance (ANOVA) followed by LSD Post Hoc tool were used to
establish statistical significance, and (P<0.05) indicated the threshold for
statistical significance. Mean and standard error of the mean (SEM) were used
to represent the values.
RESULTS
Figure 4.1: Effects of Syzygium aromaticum Extract on oestrogen level in female Wistar rats
Values represent mean ± SEM, n=3; a Significant at p<0.05 compared
to Group 1; b Significant at p<0.05 when compared to group 2; c Significant at p<0.05 when compared to group
3.
Key: Group 1: Control
Group
2: Low Dose
Group 3: Medium Dose
Group 4: High Dose
Figure 4.2: Effects
Syzygium aromaticum Extract on progesterone level in female Wistar rats
Values represent mean ± SEM, n=3; a Significant at p<0.05 compared
to Group 1; b Significant at p<0.05 when compared to group 2; c Significant at p<0.05 when compared to group
3.
Key: Group 1: Control
Group
2: Low Dose
Group 3: Medium Dose
Group 4: High Dose
Figure 4.3: Effects of Syzygium aromaticum extract on superoxide dismutase (SOD) level in female
Wistar rats
Values represent mean ± SEM, n=3; a Significant at p<0.05 compared
to Group 1; b Significant at p<0.05 when compared
to group 2; c Significant at p<0.05 when compared to
group 3.
Key: Group 1: Control
Group
2: Low Dose
Group 3: Medium Dose
Group 4: High Dose
Figure 4.4: Effects of Syzygium aromaticum extract on malondialdehyde (MAD) level in female Wistar rats
Values represent
mean ± SEM, n=3; a Significant at p<0.05 compared
to Group 1; b Significant at p<0.05 when compared to group 2; c Significant at p<0.05 when compared to group
3.
Key:
Group 1: Control
Group
2: Low Dose
Group 3: Medium Dose
Group 4: High Dose
Table 4.1: Effects of Syzygium aromaticum Extract
on reproductive hormone
and oxidative Stress
Markers in female Wistar rats
|
Groups and
Treatments of Study
Animals
|
Oestrogen (ng/ml)
|
Progesterone (ng/ml)
|
Superoxide (SOD)
(units/ml)
|
Malondialdehyde (MAD) (nm/ml)
|
|
Group 1: Control
|
87.66 ± 11.84
|
26.50
± 2.65
|
0.21
± 0.02
|
0.35
± 0.04
|
|
Group 2: Low Dose
|
78.67
± 10.10
|
32.96
± 1.85
|
0.31
± 0.03
|
0.47
± 0.02
|
|
Group 3: Medium Dose
|
92.00
± 10.39
|
40.56
± 10.42
|
0.39
± 0.05
|
0.44
± 0.04
|
|
Group 4: High Dose
|
134.33
± 8.25 a,
b, c
|
20.83
± 4.07 c
|
0.34
± 0.07
|
0.47
± 0.07
|
Values represent mean ± SEM, n=3; a Significant at p<0.05 compared
to Group 1; b Significant at p<0.05 when compared to group 2; c Significant at p<0.05 when compared to group
3.
DISCUSSION
The
results showed that the progesterone levels increased significantly and when it
increases it encourages reproduction
because it helps in the constriction of the uterus as well as the movement of sperm through the fallopian tube and
it plays a role during the normal menstruation cycle. According to (Jessie and Micheal,
2023) progesterone plays a vital role in the maintenance of the uterus during pregnancy. As such, an important issue of concern regarding the loss of
this steroid hormone correlates with miscarriage and pre- term labor. Progesterone throughout pregnancy is to maintain
and decrease levels
of vascular tone in the myometrium; it also influences the production of inflammatory makers (Khalid et al.,
2023). Also, the estrogen levels
increased significant. When estrogen
levels increase in the female reproductive system, several physiological
changes occur. Estrogen is a hormone
that is primarily responsible for the development and regulation of the female
reproductive system. One of the main effects
of increased estrogen
levels is the thickening of the uterine
lining, known as the endometrium. This prepares the uterus for possible implantation of a fertilized egg. Additionally, estrogen also stimulates the
growth and development of the ovarian
follicles, which contain the eggs, and triggers the release of luteinizing
hormone (LH), which is necessary for ovulation to occur (Mahmood, 2015).
In the menstrual cycle, estrogen
levels rise during the first
half, known as the follicular phase, and peak just before ovulation. This increase
in estrogen levels
leads to the maturation and release of an egg from the ovary (Kumar
et al., 2007). Furthermore, estrogen also plays a role in the regulation of secondary sexual characteristics such as breast
development, body hair distribution, and regulation of the menstrual cycle (Kumar et al., 2007).
Another study on female rats reported that treatment with clove extract
increased the weight of the ovary and uterus indicating estrogenic effect (Suleyman et al.,
2015; Siddiqui et al, 2011).
For the oxidative
stress markers according
to the results shown above they were significantly low, so
this simply means that cloves actually serve as a good antioxidants and it also
helps to cause stability and maintain
the pregnancy or fertility. According to (Janka Vakovα et al., 2023) oxidative stress (OS) has an important role in female
reproduction, whether it is ovulation, endometrium decasualization, menstruation, oocyte
fertilization, or development and implantation of an embryo
in the uterus. Antioxidants are very crucial for proper female
reproductive functions. They play a part
in the metabolism of Oocytes, in endometrium maturation via Nrf2 and NF - kB, and in the hormonal
regulation of vascular actions. Antioxidants can directly scavenge radicals and
act as a cofactor of highly
valuable enzymes of cell differentiation and development, or enhance the activity
of antioxidants enzymes (Janka Vaskova et al., 2023).
The datas on Figure 4.1, 4.2, 4.3 and 4.4
shows that the oestrogen level of the treated groups
were seen to be significantly (P<0.05) elevated in Group 4 (treated with high dose) when compared
to groups 1, 2 and 3 respectively. Amongst the treated groups, the pattern
of variation was observed to be dose-dependent.
The variation in progesterone levels in the
treated rats were marginally (P>0.05) different when compared to that of Group 1 (untreated/control group). However, group 4 (treated
with high dose)
was found to have progesterone level that was significantly (P<0.05) lower when compared
to that of group 3 (treated with medium dose).
Considering the changes in super oxide
dismutase (SOD) and malondialdehyde (MAD) levels across the treated rats, non-statistically
significant (P>0.05) elevations
were observed when compared to those of their respective
control groups, but these elevations did follow any uniform pattern.
CONCLUSION
The present
study revealed that the effects
of the treatment on reproductive hormone and oxidative stress markers in female Wistar rats
were dose-dependent. The high dose treatment resulted in significant elevations in oestrogen
levels and a significant decrease in progesterone levels when compared to the other treatment groups. However,
there were no statistically significant changes in superoxide dismutase (SOD) and malondialdehyde
(MAD) levels across the treated rats when compared
to their respective control groups.
ACKNOWLEDGEMENT
We
acknowledge Dr.T.M .Bekinbo
and Mrs. Precious Wadioni for their priceless
support.
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