Hindawi

Journal of Toxicology
Volume 2022, Article ID 5209136, 8 pages
https://doi.org/10.1155/2022/5209136

Research Article
Prenatal Developmental Toxicity and Histopathological
Changes of the Placenta Induced by Syzygium guineense Leaf
Extract in Rats

Melese Shenkut Abebe ,1 Kaleab Asres,2 Yonas Bekuretsion,3 Samuel Woldekidan,4

Bihonegn Sisay,4 and Girma Seyoum 5
1
Department of Anatomy, School of Medicine, College of Medicine and Health Science, Wollo University, Dessie, Ethiopia
2
Department Pharmaceutical Chemistry and Pharmacognosy, College of Health Sciences, Addis Ababa University,
Addis Ababa, Ethiopia
3
Department of Pathology, College of Health Sciences, Addis Ababa University, Addis Ababa, Ethiopia
4
Traditional and Modern Medicine Research Directorate, Ethiopian Public Health Institute, Addis Ababa, Ethiopia
5
Department of Anatomy, College of Health Sciences, Addis Ababa University, Addis Ababa, Ethiopia

Correspondence should be addressed to Melese Shenkut Abebe; melese19@yahoo.com

Received 29 April 2022; Accepted 7 October 2022; Published 11 October 2022

Academic Editor: Robyn Tanguay

Copyright © 2022 Melese Shenkut Abebe et al. This is an open access article distributed under the Creative Commons Attribution
License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is
properly cited.
Many of the traditional herbal products are served to the consumer without proper efficacy and safety investigations. A laboratory-
based experimental study was employed to investigate the toxic effects of Syzygium guineense leaf extract on the fetal development
and histopathology of the placenta in rats. Fifty pregnant Wistar albino rats were randomly allocated into five groups, each
consisting of 10 rats. S. guineense leaf extract, at doses of 250, 500, and 1000 mg/kg of body weight, was respectively administered
to groups I-III rats. Groups four and five were control and ad libitum control, respectively. The number of resorptions, im-
plantation sites, and live or dead fetuses was counted. The weight and crown-rump length of the fetuses were measured. The
histopathological investigation of the placenta was conducted. Administration of 70% ethanol extract of S. guineense leaves
reduced weight gain and food intake of pregnant rats at p value <0.05. The crown-rump length of the near-term rat fetus was
significantly reduced in rats treated with 1000 mg/kg body weight of S. guineense extract (p value <0.05). The plant extract did not
affect the number of implantations, fetal resorptions, live births, and stillbirths. The weight of the fetuses and the placentae also
decreased dose-dependently. Decidual cystic degeneration was the most prevalent histopathological change observed in a rat’s
placenta treated with 1000 mg/kg body weight of S. guineense extract. Consumption of S. guineense leaves, especially at a high dose,
may affect fetal development. Therefore, liberal use of S. guineense leaves during pregnancy should be avoided.

1. Introduction Polyphenols, essential oils, alkaloids, terpenoids, anthra-

quinones, flavonoids, tannins, saponins, glycosides, and
Syzygium guineense wall. is a commonly used medicinal triterpenes are commonly identified secondary metabolites
plant in many African countries, including Ethiopia, Eritrea, of the leaves of S. guineense [6, 7].
and Somalia [1]. Researchers reported that the plant has Many traditional herbal products are sold to consumers
demonstrated efficacy against hypertension [2], hypergly- without proper efficacy and safety evaluations [8]. This ac-
cemia [3], cancer [4], pain and inflammation [5], and many tivity has been reported in different countries. However,
others. The effectiveness of S. guineense against a variety of research should be conducted to investigate the noxious ef-
illnesses is credited to its phytochemical components. fects of herbal products antecedent to their use by consumers.
2 Journal of Toxicology

Because various undesirable results, including death, have relative humidity of 50 ± 10% under a controlled alternating
been reported from the use of herbal products [9, 10]. 12-hour light-dark cycle. The experimental rats were fed a
Prenatal exposure to chemicals may lead to abnormal standard laboratory diet and served with drinking water
development in the embryo/fetus [11, 12]. This can be unlimitedly.
manifested by the presence of malformed organs, devel-
opmental delay, functional deterioration, complete or partial
2.3. Experimental Design. The rats were mated by placing a
agenesis of organs, and fetal death. Chemical exposure
single female rat in a cage containing one unrelated male rat
during pregnancy can directly affect fetal development in-
of proven fertility. Following an overnight mating period,
dependent of their toxicity to the mother [13]. Following
the next morning, mated female rats were inspected for the
maternal exposure to chemical agents, many of these
presence of a copulatory plug. Thereupon, a vaginal smear
chemicals can pass through the placental membrane and
test was conducted, and the presence of sperm in the vaginal
reach the embryo/fetus. These chemicals have the potential
smear was considered day one of pregnancy. This study
to disrupt the normal development of their offspring [14].
employed three treatment and two control groups of rats.
Toxicological screening of any potential drug for its
Fifty pregnant rats were randomly (using a computer-based
effect on intrauterine development has become mandatory
random order generator) assigned into five groups, each
before use in humans. Investigating the hurtful effect of plant
consisting of 10 rats. The treatment groups (groups one, two,
products on intrauterine development is one of the most
and three) were treated with 250, 500, and 1000 mg/kg of the
pompous investigations to be performed [15]. In our pre-
S. guineense extract dissolved in distilled water via intra-
vious study conducted to evaluate the effect of S. guineense
gastric tube, respectively. The first control group (group IV)
on skeletal and soft tissue development, there was no sig-
was given distilled water at 1 ml/100 g of body weight. This
nificant developmental delay observed in the genital or other
group was used to ascertain whether the outcomes were due
organs [16]. On the other hand, another study reported that
to manipulation (stress) or not. The second control group
consumption of S. guineense extract did not produce sig-
(group V or ad libitum control or untouched control) was
nificant developmental anomalies in the reproductive or-
neither an extract nor distilled water treated and used as a
gans, including anogenital distance and nipple retention
standard comparison group. The treatment doses were se-
[17]. The detailed effect of S. guineense on fetal development
lected based on a previous efficacy study report [3]. The
has not been determined yet. Therefore, our study was
duration of the treatment period was day-6 through day-12
concerned with investigating the toxic effects of S. guineense
of pregnancy, which is a critical phase of organ formation in
on the fetal development and histopathology of the placenta
rats. Throughout the treatment period, the food intake of
in rats.
pregnant dams was measured daily. Pregnant rats were
weighed at the confirmation of pregnancy, beginning of
2. Materials and Methods treatment (6th day of pregnancy), end of treatment (12th day
of pregnancy), and necropsy (20th day of pregnancy).
2.1. Collection and Extraction of the Plant Material. The Weight gain at the time of gestation was computed and
leaves of S. guineense were harvested from the surroundings compared between the treatment and control groups. We
of Woliso town, 113 km away from Addis Ababa, the capital followed the methods described by Seyoum and Persaud [20]
city of Ethiopia. The test plant was verified by a taxonomist and Seyoum [21].
in the National Herbarium of Ethiopia with a voucher On day 20 of gestation, the pregnant rats were anes-
sample (MS 001) deposited for later remark. Regarding the thetized by an intraperitoneal injection of pentobarbital
extraction procedure, we followed the methods of Abebe (150 mg/kg body weight) [22] and sacrificed humanely. The
et al. [18]. Briefly, the leaves of the test plant were cleaned, abdomen was opened by a longitudinal incision and the
shade dried, broken into pieces by hand, and roughly ground gravid uterus was kept intact, and the following evaluations
by an electric mill. For extraction, the powder of the dried were carried out. The number of implantation sites was
leaves was macerated in 70% ethanol for 24 hours with counted. The number of fetal deaths and viable fetuses were
frequent rotation by an orbital shaker. After that, the blend counted by applying gentle pressure to the fetus. The
was refined by Whatman paper (No. 1, 18 cm in diameter). number and degree of resorption (early or late) sites was
The blend was evaporated with a rotatory evaporator at 40°C counted by checking uterine nodules that were not occupied
and the remaining extract was further seared in a hot water by living or recently dead fetuses. After the aforementioned
bath at 45°C. The dried (solvent-free) extract was stored in a examinations, the two horns of the uterus were incised along
refrigerator at −4°C until administration to the rats [19]. the antimesometrial border. Then, the fetuses, placentae, and
associated fetal membranes were exposed. The placentae and
fetal membranes were removed and measured separately.
2.2. Test Animals. Our test animals were Nulliparous Wistar
Fetal sex was identified as well as the weight and crown-
albino rats, 220–240 g in weight, and aged 10–12 weeks. The
rump length (CRL) of each fetus were recorded.
experimental animals were purchased from the Ethiopian
Public Health Institute (EPHI) animal rearing house. The
test animals were adapted to the new laboratory conditions 2.4. Examination of the Placenta. Each placenta was ex-
for five consecutive days. The rats were kept in a comfortable amined for any gross morphological abnormalities. Pla-
stainless-steel cage at room temperature (23 ± 3°C) with a centae of two-three fetuses/dam/group were selected for
Journal of Toxicology 3

further histopathological examination. A 3–4 mm section 3. Results

was sampled from each placenta and immersed in 10%
formalin for fixation. With regard to tissue processing and 3.1. Maternal Food Intake and Weight Gain. The levels of
staining, we followed the methods of Abebe et al. [18]. food intake are presented in Table 1. The high dose (1000 mg/
Following an overnight fixation, the tissues were dehydrated kg) treated group consumed a significantly lower amount of
by an ascending series of alcohol (40%, 50%, 70%, 80%, 90%, food compared with the controls and low dose (250 mg/kg)
and 100%). The tissues were then cleared by xylene (I, II, and treated groups. The weight gain of pregnant rats was
III). After clearing, the tissues were impregnated with melted measured between pregnancy days 6–12 and 13–20. All
paraffin wax (I and II). Finally, each sample tissue was placed treated groups showed significantly reduced weight gain
in an embedding cassette and filled with melted wax. A five during the first 6–12 days of pregnancy, compared with the
µm section was made for every block, and the ribbon was ad libitum control group. In addition, rats that received
placed on the frosted slide and kept in a hot oven (40–45°C) S. guineense extract at a 1000 mg/kg dose showed a signif-
for 20–30 minutes [23]. icant weight gain reduction during days 13–20 of pregnancy
The staining of the tissues was based on the following as compared to the control and ad libitum control groups
procedures: the slides were dewaxed with three-steps xylene (Table 1).
for five minutes in each, rehydrated with a descending series
of alcohol (absolute alcohol I, absolute alcohol II, 90% al- 3.2. Pregnancy Outcomes. In the current study, the preg-
cohol, 80% alcohol, and 70% alcohol) for two minutes in nancy outcomes measured were the number of implantation
each, washed with running tap water, followed by staining sites, resorptions, live/dead fetuses, and sex of the fetus
with Harris hematoxylin for 5–10 minutes, cleaned with (Table 2). Treatment of pregnant rats with S. guineense
running tap water for 10 minutes, immersed in acid alcohol extract did not significantly alter the average number of
for 2–3 seconds, and counterstained with eosin Y for 1–2 either implantation or resorption sites (Figure 1). In addi-
minutes. Once the slides were stained, they were subjected to tion, no significant change in the number of live or dead
an ascending series of alcohol (80%, 95%, absolute alcohol I fetuses was observed between the treatment and the control
and II) for dehydration and xylene for clearing. Finally, the groups.
cleared slides were mounted with Dibutylphthalate Poly-
styrene Xylene (DPX) and covered with a cover-slip [23].
In the stained slides, a senior pathologist investigated the 3.3. Fetal Growth. Regarding the fetal growth indices (CRL,
structural integrity of the placenta using a binocular light fetal weight, and placental weight), a significant reduction of
microscope. The decidual zone, the labyrinthine zone, giant CRL was observed in fetuses treated with 1000 mg/kg body
cells, and trophoblasts of the placenta were investigated, and weight of the plant extract, compared with the control and
important findings were photographed by an automated ad libitum control groups. The CRL of fetuses in the high
built-in digital microscope camera (Leica EC4, Germany) dose and ad libitum control groups was 5.0 ± 0.4 and
under 10× and 20× objective lens magnification. 5.7 ± 0.4, respectively. The weight of the fetuses and pla-
centae showed a dose-dependent reduction in the treatment
groups. However, this was not significant when compared to
the control or ad libitum control groups (Table 3).
2.5. Statistical Analysis. Data analysis were performed by a
statistical package for social science (SPSS) version 24. The
difference between groups was analyzed by one-way analysis 3.4. Placental Histopathology. Microscopic examination of
of variance (ANOVA) followed by Turkey post-hoc test for the placenta showed some structural changes in the decidual
multiple comparisons. Moreover, the frequency of placental and labyrinthine zones of the placenta (Figures 2 & 3).
histopathological changes was calculated and the difference Decidual cystic degeneration, hemorrhage, and trophoblast
between groups in the frequency of placental changes was proliferation were the histopathological changes observed.
checked by a chi-square test. Results are expressed as Although the frequency of decidual cystic degeneration was
mean ± standard deviation of mean (SDM) and percentages. higher in rats treated with a high dose of the plant extract,
A cutoff point value for statistical significance was p value none of the above changes were statistically significant
<0.05. (Table 4).

4. Discussion
2.6. Ethical Approval. This research was conducted follow- Prenatal development can be divided into three phases: pre-
ing an ethical approval letter obtained from the Institutional embryonic, embryonic, and fetal periods [25]. The embry-
Review Board at the College of Health Sciences, Addis Ababa onic period, day 6–12 of gestation in rats, is a critical period
University. All protocols were conducted in obedience to the where organs of the embryo can be damaged if exposed to a
recommendations of the Organization for Economic Co- teratogen [21]. In intrauterine life, the developing fetus is not
operation and Development (OECD) guidelines [24] and the fully protected from toxicants. Studies have reported that
highest standards of the laboratory of EPHI. many environmental chemicals can pass through the
4 Journal of Toxicology

Table 1: Maternal weight gain and food intake of pregnant rats treated with 70% ethanol leaf extract of Syzygium guineense.
Group
Weight gain and food intake Group V Ad- libitum
Group I 250 mg/kg Group II 500 mg/kg Group III 1000 mg/kg Group IV control
control
Food intake/day (g) n � 10 199.2 ± 4.5 191.4 ± 8.0 ∗∗ 180.0 ± 12.1∗ ! 196.6 ± 10.0 214.0 ± 4.5∗
Maternal weight gain/dam (g)
Day 6–12 17.8 ± 8.5 17.2 ± 7.4 15.8 ± 7.5 19.0 ± 12.5 40.2 ± 5.8∗
Day 13–20 69.7 ± 3.4 67.2 ± 6.6 60.5 ± 5.6 ∗∗∗ 72.2 ± 5.6 72.8 ± 9.6
The results are expressed as mean ± SDM, ∗ significant difference with all the other groups, ∗ ! significant difference with Group I, IV and V, ∗∗
significant
difference with group V, ∗ ∗ ∗ significant difference with group IV and V (for all p value <0.05); and one-way ANOVA.

Table 2: Pregnancy outcomes of rats treated with 70% ethanol leaf extract of Syzygium guineense.
Group
Variables
250 mg/kg n � 10 500 mg/kg n � 10 1000 mg/kg n � 10 Control n � 10 Ad libitum control n � 10
Number of fetuses 93 91 88 95 115
Number of implantation sites/litter 9.8 ± 2.1 10.0 ± 2.5 10.0 ± 1.4 10.5 ± 0.5 11.7 ± 1.2
Number of resorption sites/litter 0.5 ± 0.8 0.9 ± 0.9 1.0 ± 0.9 1.0 ± 1.6 0.2 ± 0.4
Live fetuses/litter 9.3 ± 2.7 9.1 ± 3.5 8.8 ± 1.6 9.5 ± 1.9 11.5 ± 1.4
Dead fetuses/litter 0 0 0.2 ± 0.4 0 0
Number of male fetuses/dam 4.5 ± 1.6 4.3 ± 1.2 3.7 ± 1.5 5.0 ± 1.1 5.3 ± 1.2
Number of female fetuses/dam 4.8 ± 2.7 4.8 ± 2.4 5.3 ± 2.3 4.5 ± 2.4 6.2 ± 0.8
The results are expressed as mean ± SDM, One-Way ANOVA; n: number of dams.

*
*
* *
*

(a) (b) (c) (d) (e)

Figure 1: Gravid uterus of rat from group I, (a) group II, (b) group III, (c) group IV, and (d) group V (e) indicating implantation sites (∗)
and fetal resorption (arrow) following administration of ethanol leaf extract of S. guineense.

Table 3: Fetal growth indices of rat fetuses following the administration of 70% ethanol leaf extract of Syzygium guineense.
Group
Variables
250 mg/kg n � 10 500 mg/kg n � 10 1000 mg/kg n � 10 Control n � 10 Ad libitum control n � 10
CRL/fetus (cm) 5.3 ± 0.2 5.2 ± 0.4 5.0 ± 0.4∗ 5.5 ± 0.2 5.7 ± 0.4
Fetal weight (g) 5.8 ± 0.9 5.1 ± 0.5 4.9 ± 0.5 5.5 ± 0.6 5.1 ± 0.8
Placental weight (g) 0.7 ± 0.2 0.6 ± 0.1 0.6 ± 0.1 0.7 ± 0.1 0.6 ± 0.05
The results are expressed as mean ± SDM, ∗ significant difference with control and ad libitum control groups (p value <0.05), One-Way ANOVA; CRL:
Crown-rump length.

placental membrane, and as many as 200 foreign chemicals The objective of the present study was to evaluate the
have been detected in the blood samples taken from the toxic effects of prenatal exposure of S. guineense on the fetal
umbilical cord [26]. Due to the undeveloped metabolic development and histopathology of the placenta. In the
function of the liver and the excretion capacity of the kidney, present study, administration of the 70% ethanol extract
the level of toxicants in the fetal circulation is much greater S. guineense to the pregnant rats during the crucial period of
than in the maternal circulation [27]. Toxic agents can di- organogenesis (day-6 through day-12 of gestation) resulted
rectly or indirectly affect embryos/fetuses. When the toxic in reduced food intake and weight gain of the pregnant dams
agents cross the placental membrane, they directly damage and CRL of 20-day old rat fetuses in the high dose (1000 mg/
the developing embryonic/fetal tissue. Indirectly, toxicants kg body weight) treated group.
that damage the placental tissue and compromise placental Maternal weight gain during the treatment period was
function might impede the development of embryos/fetuses affected by the administration of the plant extract despite
[28]. similar intake of food. It was witnessed by a significant and
Journal of Toxicology 5

DB
BV

TZ

LZ

(a) (b) (c)

(d)

Figure 2: Photomicrograph of rat placenta of group IV, (a) group V, and (b) group I (c) showing normal architecture of the placenta and
group II (d) showing hematoma in the different zones of the placenta (∗); DB: Decidua basalis, TZ: Trophoblastic zone, LZ: Labyrinthine
zone, BV: Blood vessel; H and E stain, 100× total magnification.

(a) (b)

Figure 3: Photomicrograph of rat placenta (a & b) from a rat treated with 1000 mg/kg of ethanol extract of S. guineense leaves (group III)
showing decidual cystic degeneration (∗) and trophoblast proliferation (arrow); H and E stain, 200× total magnification.

Table 4: Microscopic placental abnormalities of rats following administration of 70% ethanol leaf extracts of Syzygium guineense.
Percent of placental abnormalities
Group
Decidual cystic degeneration Hemorrhage/hematoma Trophoblast proliferation
Group I (250 mg/kg) 0 0 0
Group II (500 mg/kg) 10 10 10
Group III (1000 mg/kg) 20 0 10
Group IV (control) 10 10 0
Group V (Ad libitum control) 0 0 0
Results are expressed as percentage of placental abnormalities, Chi-square.
6 Journal of Toxicology

dose-dependent reduction in weight gain in the rats treated contains newly formed vasculatures. The decidua basalis
with higher doses of the test plant. Reduction in weight gain mainly develops during early pregnancy, and it undergoes
was also reported by other researchers: Abba et al. [29], Loha regression after gestational day 11 in rats. As a result, de-
et al. [30], and Amare [31], who conducted toxicity studies cidua is less sensitive to chemical exposure in the em-
on nonpregnant rats and mice following administration of bryogenesis period than the other parts of the placenta.
S. guineense extract. Moreover, the food intake of pregnant Moreover, hemorrhage can be seen when decidual regres-
rats in the treatment group decreased significantly compared sion occurs [45]. This might be the justification for the
to the control groups. A study conducted by Rogers and presence of decidual cystic degeneration and hemorrhage in
Kavlock [32] reported that decreased food intake can induce both treated and control groups. On the other hand,
weight loss and other clinical signs. Therefore, this could be spongiotrophoblasts, glycogen cells, and trophoblastic giant
the reason for the decrease in weight gain during gestation. cells were not noticeably affected by treatment with the plant
As reported by Chung et al. [33], tannins damage the epi- extract.
thelial coverings of the digestive tract and reduce food in-
take. S. guineense is a tannin-rich plant [34, 35], and the 5. Limitations of the Study
presence of tannins in high concentrations may explain the
reduced food intake in the S. guineense treated rats. The current study provided evidence regarding the devel-
Retarded fetal development in vivo is manifested by opmental toxicity of S. guineense, which contributes to the
decreased fetal weight, placental weight, and CRL [36]. In knowledge of science in terms of the developmental toxicity
the current experiment, treatment with 1000 mg/kg of of S. guineense. However, it is not without limitations. The
S. guineense extract significantly reduced the CRL of the main limitation is that, due to financial constraints, ad-
fetuses when compared with the control groups. In addition, vanced tests such as immunohistochemistry and electron
there was a reduction in fetal and placental weights. This microscopy were not included. Secondly, the test substance
finding is in line with the previous study [16]. The leaves of was administered only from day 6–12 of gestation. It would
S. guineense possess secondary metabolites such as terpe- have had value if the treatment period was the whole period
noids, anthraquinones, flavonoids, tannins, saponins, gly- of pregnancy.
cosides, triterpenes, and phenols that pass through the
placental membrane and hurt fetal development [7, 37, 38]. 6. Conclusion
These secondary metabolites of S. guineense might be re-
In conclusion, administration of 1000 mg/kg body weight of
sponsible for the decrease in the CRL of the treated rat
70% ethanol extract of S. guineense leaf delayed fetal de-
fetuses [16].
velopment as witnessed by reduced CRL of 20-day old rat
The plant extract had no effect on the other pregnancy
fetuses. High dose treatment with S. guineense extract also
outcomes, including the number of implantations, fetal
decreased food intake and weight gain of the pregnant rats
resorptions, live births, and stillbirths. This refers that the
that showed its toxicity at high dose. Therefore, care should
test plant may not have a significant detrimental effect on the
be taken while consuming the S. guineense leaf during
progression of pregnancy in rats.
pregnancy, especially in a high dose. The results of our study
The placenta plays a central role in the transfer of nu-
would be a benchmark for further investigation of the test
trients, gases, metabolic wastes, drugs, and immunoglobu-
plant and for incorporating the plant into modern phar-
lins between the mother and the embryo/fetus. It also allows
maceutical products. Further studies should be conducted by
the passage of toxicants, mycotoxins, plant alkaloids, and
administering the plant extract during the whole period of
many others [39–42]. Due to its function, the placenta is
gestation and isolating secondary metabolites of S. guineense
highly sensitive to toxins [39]. Succeeding maternal expo-
leaf with serum level determination to identify the toxic
sure to chemicals, many of these chemicals can pass through
nature and mechanism of action. Based on these findings, it
the placental membrane and reach the embryo/fetus. Thus,
is also advisable to investigate the clinical applicability of
the chemicals can affect the fetus, the placenta, and even the
S. guineense.
mother herself [14]. Administration of chemical agents in
the early embryonic period, since the trophoblast cell dif-
ferentiation is not complete to make the placental membrane
Abbreviations
[43], can affect the development of embryonic tissue and the ANOVA: One-way analysis of variance
placenta [44]. CRL: Crown-rump length
In the present study, decidual cystic degeneration, DPX: Dibutylphthalate polystyrene xylene
hemorrhage, and trophoblast proliferation were seen in both EPHI: Ethiopian public health institute
treatment and control groups. A greater frequency of de- OECD: Organization for economic co-operation and
cidual cystic degeneration was prevalent in rats treated with development
1000 mg/kg body weight of S. guineense. However, these SDM: Standard deviation of mean
alterations in the microscopic structures of the placenta were SPSS: Statistical package for social science.
not statistically significant. Similarly, in another study, there
were no significant histological alterations observed in the Data Availability
liver and kidney of rats treated with S. guineense extract [18].
The decidua basalis is found at the base of the placenta and it All data are included in the manuscript.
Journal of Toxicology 7

Conflicts of Interest [8] W. M. Bandaranayake, “Quality control, screening, toxicity,

and regulation of herbal drugs,” in Modern Phytomedicine:
All authors declare that they have no conflicts of interest. Turning Medicinal Plants into DrugsWiley-VCH Verlag
GmbH & Co, Berlin, Germany, 2006.
[9] J.-P. Cosyns, M. Jadoul, J.-P. Squifflet, F.-X. Wese, and
Authors’ Contributions C. van Ypersele de Strihou, “Urothelial lesions in Chinese-
MSA developed the proposal and collected the data, ana- herb nephropathy,” American Journal of Kidney Diseases,
vol. 33, no. 6, pp. 1011–1017, 1999.
lyzed and interpreted the results, and was the major con-
[10] E. Ernst, “Toxic heavy metals and undeclared drugs in Asian
tributor in writing the mother document and the
herbal medicines,” Trends in Pharmacological Sciences, vol. 23,
manuscript. KA, YB, SW, BS, and SG critically reviewed both no. 3, pp. 136–139, 2002.
the proposal and study and contributed to the study design [11] M. Erdemli, Y. Turkoz, E. Altinoz, E. Elibol, and Z. Dogan,
development and interpretation of the study. YB performed “Investigation of the effects of acrylamide applied during
identification and interpretation of histopathological find- pregnancy on fetal brain development in rats and protective
ings on placenta sections. GS conceived and supervised the role of the vitamin E,” Human & Experimental Toxicology,
entire research and masterminded the research protocol, vol. 35, no. 12, pp. 1337–1344, 2016.
including the design. All authors read and approved the final [12] M. E. Erdemli, M. Arif Aladag, E. Altinoz et al., “Acrylamide
manuscript. applied during pregnancy causes the neurotoxic effect by
lowering BDNF levels in the fetal brain,” Neurotoxicology and
Teratology, vol. 67, pp. 37–43, 2018.
Acknowledgments [13] K. Khera, H. C. Grice, and D. Clegg, Interpretation and Ex-
Even though they do not have a role in writing or publishing trapolation of Reproductive Data to Establish Human Safety
the manuscript, EPHI and Addis Ababa University provided Standards, Springer Science & Business Media, Berlin, Ger-
many, 2012.
support to our experiment. This manuscript is based on a
[14] R. K. Gupta and R. C. Gupta, Placental Toxicity. Reproductive
PhD dissertation of Melese Shenkut Abebe submitted to
and Developmental Toxicology, Elsevier, Amsterdam, Neth-
Addis Ababa University, School of Graduate Studies, College erlands, 2022.
of Health Sciences, Department of Anatomy [17]. Addis [15] K. Augustine-Rauch, C. X. Zhang, and J. M. Panzica-Kelly, “A
Ababa University and Ethiopian Public Health Institute are developmental toxicology assay platform for screening tera-
duly acknowledged for their contribution to the success of togenic liability of pharmaceutical compounds,” Birth Defects
this work. Research Part B: Developmental and Reproductive Toxicology,
vol. 107, no. 1, pp. 4–20, 2016.
References [16] M. Abebe, K. Asres, Y. Bekuretsion, S. Woldkidan, E. Debebe,
and G. Seyoum, “Teratogenic effect of high dose of Syzygium
[1] J. Nvau, P. Oladosu, and A. Orishadipe, “Antimycobacterial guineense (myrtaceae) leaves on wistar albino rat embryos
evaluation of some medicinal plants used in Plateau State of and fetuses,” Evidence-based Complementary and Alternative
Nigeria for the treatment of tuberculosis,” Agriculture and Medicine, vol. 2021, Article ID 6677395, 10 pages, 2021.
Biology Journal of North America, vol. 2, no. 9, pp. 1270–1272, [17] M. S. Abebe, “Extended one-generation reproductive toxicity
2011. and teratogenicity of ethanol leaf extract of Syzygium gui-
[2] Y. Ayele, K. Urga, and E. Engidawork, “Evaluation of in vivo neense Wall,” in RatsAddis Ababa University, Addis Ababa,
antihypertensive and in vitro vasodepressor activities of the Ethiopia, 2021.
leaf extract of Syzygium guineense (Willd) DC,” Phytotherapy [18] M. S. Abebe, K. Asres, Y. Bekuretsion, A. Abebe, D. Bikila, and
Research, vol. 24, no. 10, pp. 1457–1462, 2010. G. Seyoum, “Sub-chronic toxicity of ethanol leaf extract of
[3] I. C. Ezenyi, O. N. Mbamalu, L. Balogun, L. Omorogbe, Syzygium guineense on the biochemical parameters and
F. S. Ameh, and O. A. Salawu, “Antidiabetic potentials of histopathology of liver and kidney in the rats,” Toxicology
Syzygium guineense methanol leaf extract,” Journal of Phy- Reports, vol. 8, pp. 822–828, 2021.
topharmacology, vol. 5, no. 4, pp. 150–156, 2016. [19] Q.-W. Zhang, L.-G. Lin, and W.-C. Ye, “Techniques for ex-
[4] A. Koval, C. A. Pieme, E. F. Queiroz et al., “Tannins from
traction and isolation of natural products: a comprehensive
Syzygium guineense suppress Wnt signaling and proliferation
review,” Chinese Medicine, vol. 13, no. 1, pp. 20–26, 2018.
of Wnt-dependent tumors through a direct effect on secreted
[20] G. Seyoum and T. Persaud, “Protective influence of zinc
Wnts,” Cancer letters, vol. 435, pp. 110–120, 2018.
against the deleterious effects of ethanol in postimplantation
[5] L. D. Ior, I. Otimenyin, and M. Umar, “Anti-inflammatory
and analgesic activities of the ethanolic extract of the leaf of rat embryos in vivo,” Experimental & Toxicologic Pathology,
Syzygium guineense in rats and mice,” IOSR Journal of vol. 47, no. 1, pp. 75–79, 1995.
Pharmacy, vol. 2, no. 4, pp. 33–36, 2012. [21] G. Seyoum, “Influence of methionine supplementation on
[6] J. Djoukeng, E. Abou-Mansour, Rl Tabacchi, A. Tapondjou, nicotine teratogenicity in the rat,” Ethiopian Pharmaceutical
H. Bouda, and D. Lontsi, “Antibacterial triterpenes from Journal, vol. 32, no. 1, pp. 37–54, 2017.
Syzygium guineense (myrtaceae),” Journal of Ethno- [22] W. Underwood and R. Anthony, AVMA Guidelines for the
pharmacology, vol. 101, no. 3, pp. 283–286, 2005. Euthanasia of Animals: 2020 Edition, American Veterinary
[7] S. A. Tadesse and Z. B. Wubneh, “Antimalarial activity of Medical Association, Schaumburg, Illinois, USA, 2020.
Syzygium guineense during early and established Plasmo- [23] J. D. Bancroft, A. D. Floyd, and S. K. Suvarna, Bancroft’s
dium infection in rodent models,” BMC Complementary and Theory and Practice of Histological Techniques, Cherchil
Alternative Medicine, vol. 17, no. 1, p. 21, 2017. Livingstone, London, UK, 2013.
8 Journal of Toxicology

[24] OECD, OECD Guideline for Testing of Chemicals. Prenatal [40] N. M. Gude, C. T. Roberts, B. Kalionis, and R. G. King,
Developmental Toxicity Study. Test No. 414, OECD, Mexico, “Growth and function of the normal human placenta,”
2018. Thrombosis Research, vol. 114, no. 5-6, pp. 397–407, 2004.
[25] K. L. Moore, T. V. N. Persaud, and M. G. Torchia, The De- [41] G. T. Knipp, K. L. Audus, and M. J. Soares, “Nutrient
veloping Human: Clinically Oriented Embryology, Elsevier transport across the placenta,” Advanced Drug Delivery Re-
Health Sciences, Amsterdam, Netherlands, 2018. views, vol. 38, no. 1, pp. 41–58, 1999.
[26] L. L. Needham, P. Grandjean, B. Heinzow et al., “Partition of [42] G. M. Pacifici and R. Nottoli, “Placental transfer of drugs
environmental chemicals between maternal and fetal blood administered to the mother,” Clinical Pharmacokinetics,
and tissues,” Environmental science & technology, vol. 45, vol. 28, no. 3, pp. 235–269, 1995.
no. 3, pp. 1121–1126, 2011. [43] S. L. Adamson, Y. Lu, K. J. Whiteley et al., “Interactions
[27] G. Ahmad, L. Halsall, and S. Bondy, “Persistence of phen- between trophoblast cells and the maternal and fetal circu-
cyclidine in fetal brain,” Brain Research, vol. 415, no. 1, lation in the mouse placenta,” Developmental biology, vol. 250,
pp. 194–196, 1987. no. 2, pp. 358–373, 2002.
[28] G. P. Daston, “Relationships between maternal and devel- [44] G. d A. Costa, T. C. Galvão, A. D. Bacchi, E. G. Moreira, and
opmental toxicity,” in Developmental ToxicologyRaven Press, M. J. S. Salles, “Investigation of possible teratogenic effects in
New York, NY, USA, 1994. the offspring of mice exposed to methylphenidate during
[29] S. Abba, O. O Dare, M. Joseph, and O. Eyinna, “Hemorrhagic pregnancy,” Reproductive BioMedicine Online, vol. 32, no. 2,
centrolobar necrosis and cytoplasmic vacuolation of the he- pp. 170–177, 2016.
patocytes in Syzygium guineense chronic treated mice,” In- [45] D. Dai, B. C. Moulton, and T. F. Ogle, “Regression of the
ternational Journal of Anatomy and Applied Physiology, vol. 4, decidualized mesometrium and decidual cell apoptosis are
no. 4, pp. 99–102, 2018. associated with a shift in expression of Bcl2 family members,”
[30] M. Loha, A. Mulu, S. M. Abay, W. Ergete, and B. Geleta, Biology of Reproduction, vol. 63, no. 1, pp. 188–195, 2000.
“Acute and subacute toxicity of methanol extract of Syzygium
guineense leaves on the histology of the liver and kidney and
biochemical compositions of blood in rats,” Evidence-based
Complementary and Alternative Medicine, vol. 2019, Article
ID 5702159, 15 pages, 2019.
[31] G. Amare, Chronic Effects of the Aqueous Leaf Extract of
Syzygium Guineense on Some Blood Parameters and Histo-
pathology of Liver and Kidney of Mice, Addis Ababa Uni-
versity, Addis Ababa, Ethiopia, 2009.
[32] J. M. Rogers and R. J. Kavlock, Developmental Toxicology.
Reproductive and Developmental Toxicology, Marcel Dekker,
New York, NY, USA, 1998.
[33] K.-T. Chung, T. Y. Wong, C.-I. Wei, Y.-W. Huang, and Y. Lin,
“Tannins and human health: a review,” Critical Reviews in
Food Science and Nutrition, vol. 38, no. 6, pp. 421–464, 1998.
[34] T. L. Nguyen, A. Rusten, M. S. Bugge et al., “Flavonoids,
gallotannins and ellagitannins in Syzygium guineense and the
traditional use among Malian healers,” Journal of Ethno-
pharmacology, vol. 192, pp. 450–458, 2016.
[35] C. A. Pieme, J. Ngoupayo, C. H. K.-K. Nkoulou et al.,
“Syzyguim guineense extracts show antioxidant activities and
beneficial activities on oxidative stress induced by ferric
chloride in the liver homogenate,” Antioxidants, vol. 3, no. 3,
pp. 618–635, 2014.
[36] C. V. Chang, A. C. Felı́cio, J. E. D P. Reis, M. D O. Guerra, and
V. M. Peters, “Fetal toxicity of Solanum lycocarpum (Sol-
anaceae) in rats,” Journal of Ethnopharmacology, vol. 81, no. 2,
pp. 265–269, 2002.
[37] D. R. Doerge, M. I. Churchwell, H. C. Chang, R. R. Newbold,
and K. B. Delclos, “Placental transfer of the soy isoflavone
genistein following dietary and gavage administration to
Sprague Dawley rats,” Reproductive Toxicology, vol. 15, no. 2,
pp. 105–110, 2001.
[38] J. P. S. V. D. Elst, D. van der Heide, H. Rokos,
G. M. de Escobar, and J. Köhrle, “Synthetic flavonoids cross
the placenta in the rat and are found in fetal brain,” American
Journal of Physiology Endocrinology and Metabolism, vol. 274,
no. 2, pp. E253–E256, 1998.
[39] C. Göhner, J. Svensson-Arvelund, C. Pfarrer et al., “The
placenta in toxicology. Part IV: battery of toxicological test
systems based on human placenta,” Toxicologic Pathology,
vol. 42, no. 2, pp. 345–351, 2014.