Redox Report

Communications in Free Radical Research

ISSN: (Print) (Online) Journal homepage: www.tandfonline.com/journals/yrer20

Effect of acetogenin fraction of Annona muricata

leaves on antioxidant status and some indices of
benign prostatic hyperplasia in rats

Patience N. Ogbu, Evelyn O. Ugota, Rita U. Onwuka, Ikechukwu M. Ogbu &

Chinyere Aloke

To cite this article: Patience N. Ogbu, Evelyn O. Ugota, Rita U. Onwuka, Ikechukwu M. Ogbu &
Chinyere Aloke (2020) Effect of acetogenin fraction of Annona muricata leaves on antioxidant
status and some indices of benign prostatic hyperplasia in rats, Redox Report, 25:1, 80-86, DOI:
10.1080/13510002.2020.1804711

To link to this article: https://doi.org/10.1080/13510002.2020.1804711

© 2020 The Author(s). Published by Informa

UK Limited, trading as Taylor & Francis
Group

Published online: 02 Sep 2020.

Submit your article to this journal

Article views: 2172

View related articles

View Crossmark data

Citing articles: 7 View citing articles

Full Terms & Conditions of access and use can be found at

https://www.tandfonline.com/action/journalInformation?journalCode=yrer20
REDOX REPORT
2020, VOL. 25, NO. 1, 80–86
https://doi.org/10.1080/13510002.2020.1804711

Effect of acetogenin fraction of Annona muricata leaves on antioxidant status and

some indices of benign prostatic hyperplasia in rats
a
Patience N. Ogbu , Evelyn O. Ugotaa, Rita U. Onwukaa, Ikechukwu M. Ogbu b
and Chinyere Alokea
a
Department of Medical Biochemistry, Faculty of Basic Medical Sciences, Alex Ekwueme Federal University Ndufu-Alike, Ikwo, Nigeria; bDepartment
of Chemistry/Biochemistry, Faculty of Sciences, Alex Ekwueme Federal University Ndufu-Alike, Ikwo, Nigeria

ABSTRACT KEYWORDS
Objectives: This work investigated the effect of acetogenin-rich fraction of Annona muricata leaves Benign prostatic hyperplasia;
(AFAL) on antioxidant status and some markers of benign prostatic hyperplasia (BPH) in rats. Annona muricata; acetogenin
Methods: BPH was experimentally induced in the rats by subcutaneous injection of testosterone fraction; leaves extract;
propionate (TP, 3 mg/kg) for 28 consecutive days. The rats were administered orally different doses antioxidants
of AFAL (100 and 200 mg/kg) for 7 days. Prostate-specific antigen (PSA), prostate weight, relative
prostate weight, prostate protein content and oxidative stress indices of the rats were evaluated.
Results: It was observed that 200 mg/kg AFAL significantly reduced the PSA level, mean prostate
weights and mean relative prostate weights of the test rats compared to the TP group, and the
values were not significantly different from the normal control and group treated with a standard
drug. The plant extract also significantly enhanced the antioxidant capacity of the test rats which
were evidently compromised in the group that received the exogenous hormone alone.
Histopathology of the prostate showed a marked recovery for the test rats after treatment with AFAL.
Conclusion: Oral administration of acetogenin-rich fraction of Annona muricata leaves ameliorated
TP-induced BPH in rats and significantly enhanced the antioxidant capacity of the rats.

1. Introduction
phytochemical agents are also effective inhibitors of 5α-
Benign prostatic hyperplasia (BPH) is an age-related enlarge- reductase [8] and could contribute significantly to BPH
ment of the prostate gland due to the unregulated growth treatment.
of prostate cells [1,2]. The abnormal proliferation of prostatic Lots of side effects, such as decreased libido, erectile dys-
stromal cells leads to the formation of discrete nodules in function, dizziness, retrograde ejaculation and orthostatic
the periurethral region that brings about acute and chronic hypotension associated with the existing BPH drugs [9,10],
urinary retention, bladder outlet obstruction, urinary tract have increased interest and research activities on alternative
infection, urosepsis, bladder stones and hematuria [1,3]. BPH treatment options. The use of phytotherapy for the preven-
is a progressive disease and its prevalence increases with age. tion and treatment of BPH is gaining popularity [11] due to
Several studies have strongly linked etiology of BPH to an its promising efficacy, milder side effects and affordability
imbalance in steroid hormone metabolism, remodeling in compared to most other treatment options. Anti-BPH proper-
ageing prostate, systemic inflammation and oxidative stress ties of some plants, including saw palmetto, Pygeum africa-
associated with metabolic syndrome, among other factors num, Secale cereale and Phellodendron amurense, have been
[4,5]. The existing treatment options for BPH include validated by several scientific investigations [12–14] and are
medical therapy with α-blockers or 5α-reductase inhibitors, widely used for the prevention and treatment of the disease.
surgery and phytotherapy [2,6]. While α-blocker relaxes Annona muricata, commonly known as soursop, belongs to
smooth muscles of the prostate and the bladder neck to the Annonaceae family. The plant is widely known for its
relieve urinary obstruction caused by an enlarged prostate, anticancer properties [15]. Aside this popular medicinal use,
5α-reductase inhibitors prevent the conversion of testoster- a wide range of ethnomedicinal activities have also been
one to dihydrotestosterone (DHT), thereby leading to the attributed to different parts of the plant owing to some of
shrinkage of prostate. DHT is an active metabolic product its properties including anti-inflammatory, antiproliferative,
from the reduction of testosterone by 5α-reductase. It plays hypoglycemic, sedative, smooth muscle relaxant and anti-
a critical role in the growth of prostate by binding to the spasmodic effects [16,17]. Some indigenous communities in
nuclear androgen receptor, thereby inducing synthesis of Africa including Nigeria use A. muricata in their folk medicine.
growth factors that act on prostatic epithelia and stroma, Leaf extract of the plant is used to alleviate difficulty associ-
resulting in prostate enlargement [7]. Consequently, inhibitors ated with urination in certain communities in Eastern part of
of 5α-reductase that block the production of DHT ultimately Nigeria. Although Asare et al. [18] reported that the aqueous
slow down the development of BPH. Common inhibitors of extract of the plant leaf exhibited antiproliferative activity
5α-reductase are pharmacological agents such as dutasteride against BPH-1 cells, there is still paucity of information on
and finasteride. However, there is strong evidence that some the possible usefulness of this plant in the treatment of BPH.

CONTACT Patience N. Ogbu [email protected] Department of Medical Biochemistry, Faculty of Basic Medical Sciences, Alex Ekwueme Federal
University Ndufu-Alike, Ikwo, Ebonyi State, Nigeria
© 2020 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use,
distribution, and reproduction in any medium, provided the original work is properly cited.
 REDOX REPORT 81

It has been shown that plant-derived medications exhibit Kedde reagents. Formation of pink to red-purple color on
their anti-BPH effect through different mechanisms including addition of Kedde reagents indicated the presence of anno-
antiandrogenic, antiproliferative, anti-inflammatory and anti- naceous acetogenin [26,27]. Fractions tested positive with
oxidant activities [8,19–21]. Antioxidants are known to miti- Kedde reagents were combined and concentrated at 50°C.
gate the deleterious effect of oxidative stress - a factor The extract was exposed in 100 ml beakers for five days to
believed to play a great role in the development of age- ensure total evaporation of organic solvent before being
related diseases such as BPH [22,23]. In addition to antiproli- used for the experiment.
ferative effect, in vitro studies have indicated that It has been shown from previous studies that acetogenin
A. muricata leaf extract also exhibits a remarkable antioxidant fraction obtained by column chromatography of ethanol
activity [24,25]. Thus, investigation of the effect of this plant extract of A. muricata leaves contains predominantly anno-
material on BPH alongside its antioxidant activity could naceous acetogenins [28,29]; however, the presence of little
provide more useful information on its potential anti-BPH amount of flavonoid has equally been detected in the fraction
properties. [28].
Annonaceous acetogenins are the major constituents of
A. muricata and many pharmacological activities of the
plant have been attributed to these bioactive constituents
2.4. Acute toxicity test
[15–17]. In this study, the effect of acetogenin-rich fraction The median lethal dose (LD50) of AFAL was determined
of A. muricata leaves was investigated on testosterone propio- according to the method of Lorke [30]. Thirteen BALB/c
nate-induced BPH in Wistar rats through the measurement of albino mice weighing 20–22 g were used for the LD50. They
antioxidant indices and some other biochemical parameters were purchased from the Veterinary Medicine Department,
including prostate-specific antigen, prostate weight, relative University of Nigeria, Nsukka. The mice were kept in standard
prostate weight and prostate protein content. cages at the average ambient temperature of 26 ± 1°C, 12 h
light −12 h dark cycle and were fed commercial mice chow
and tap water ad libitum. In stage one, the animals were
2. Materials and methods placed in three different groups of three mice each and
2.1. Chemicals and drug were administered graded oral doses of 10, 100 and
1000 mg/kg body weight respectively of AFAL dissolved in a
The testosterone propionate (TP) used for the induction of
mixture of dimethyl sulfoxide (DMSO) and H2O (in the ratio
BPH was purchased from Sigma-Aldrich chemical company,
of 0.5: 9.5 v/v). Then in stage two, three mice were placed in
Germany. The enzyme-linked immunosorbent assay test kit
three different groups and were orally administered 1600,
used for PSA determination was a product of Elabscience Bio-
2900 and 5000 mg/kg body weight of the dissolved AFAL,
technology, Inc. Houston, Texas, United State. AvodartTM
while one mouse served as control. The mice were observed
(Dutasteride) was manufactured by GlaxoSmithKline Pharma-
for toxicity signs and possible death in each group within
ceuticals S.A., 189 Grunwaldzka Str, 60–322 Poznan, Poland.
24 h of administration for the lethal dose determination.
Reagents used for the assays were commercial test kits pur-
The LD50 of AFAL was obtained at 2900 mg/kg body weight
chased from Randox Laboratories Ltd., Crumlin, Antrim, UK.
which was considered safe and, based on this, doses of 100
Thiobarbituric acid (TBA) was purchased from HiMedia Lab-
and 200 mg/kg were selected for the experiment.
oratories Pvt. Ltd, Mumbai, India. All other chemicals and
reagents used for this study were of analytical grade.
2.5. Experimental design
2.2. Sample collection and extraction Twenty-five male Wistar rats weighing 110–120 g were pur-
chased from the Veterinary Medicine Department, University
Fresh green leaves of A. muricata were collected from Ikwo of Nigeria, Nsukka. They were maintained under standard
local government area, Ebonyi State, Nigeria. The plant environmental conditions and were allowed free access to
leaves were authenticated at the Department of Agriculture, commercial grower’s feed and clean water ad libitum. The
Alex Ekwueme Federal University Ndufu-Alike. They were sub- rats were acclimatized for one week before randomly distrib-
sequently washed with clean water and air-dried at room uted into five groups of five rats each.
temperature under shade. The dried leaves were milled into The study groups and treatment protocol used in this
fine powder using a milling machine. About 560 g of the experiment are shown in Table 1. BPH was induced in the
ground material was soaked in 2.5 L of ethanol (95.5%) for
72 h. The mixture was then filtered with Whatman filter
paper (grade 1) and concentrated at 50°C to about 50 mL. Table 1. Study groups and treatment protocol.
Subcutaneous
Group injection Oral administration
2.3. Fractionation of the extract Normal Olive oil (1 ml/kg) 1 ml/kg DMSO/H20 (0.5: 9.5 v/
v)
The ethanol extract was first partitioned in n-hexane (100 mL TP TP (3 mg/kg) 1 ml/kg DMSO/H20 (0.5: 9.5 v/
×3) using a separating funnel. The ethanol layer was then con- v)
TP + Dutasteride TP (3 mg/kg) Dutasteride (0.5 mg/70 kg)
centrated to obtain a solid crude extract. The crude extract TP + AFAL (100 mg/ TP (3 mg/kg) AFAL (100 mg/kg)
was subjected to column chromatography (open column, kg)
silica gel 60), using the mixture of solvents: hexane/ethyl TP + AFAL (200 mg/ TP (3 mg/kg) AFAL (200 mg/kg)
kg)
acetate (93/7 v/v), dichloromethane: ethanol (93/7, 80/20
All animals were fed rat chow and allowed free access to water throughout the
and 50/50 v/v). Eluted fractions were collected with 100 mL period of the experiment. TP = testosterone propionate, AFAL = acetogenin-
beakers and were tested for annonaceous acetogenin using rich fraction of A. muricata leaves.
82 P. N. OGBU ET AL.

rats by subcutaneous injection of testosterone propionate were subsequently viewed and photomicrographs were
(3 mg/kg body weight, dissolved in olive oil) for 28 days taken to evaluate the structural changes.
[31]. Successful induction of BPH was ascertained by testing
PSA level of the rats on the 29th day. Then oral administration
of the acetogenin-rich fraction dissolved in DMSO/H2O (0.5: 2.8. Statistical analysis
9.5 v/v) was done for one week. On the last day of the exper- Statistical analysis was performed using the Statistical
iment (day 36th), the animals were fasted, weighed using a Package for the Social Sciences software (SPSS Institute,
sensitive balance and bled by cardiac puncture. Blood Armonk, NY, U.S.A.) version 20. The results were expressed
samples were collected in plain sample bottles and centri- as mean ± standard error of mean (SEM). The level of hom-
fuged (3000 g for 15 min) for the separation of serum. The ogeneity among the groups was evaluated using one-way
prostates were excised, washed with cold saline solution, analysis of variance followed by post-hoc multiple compari-
dried with adsorbent paper and weighed on sensitive sons with the Duncan’s tests to detect the significant differ-
balance. One prostate per group was selected and a section ences between the groups at p < 0.05.
of the dorsolateral lobe was processed for histology. The
remaining prostates were used for biochemical analyses. Rela-
tive prostate weight of the rats was calculated using the 3. Results
method of Kalu et al. [10]: Relative prostate weight = (Total
3.1. Effect of the acetogenin-rich fraction on PSA level
prostate weight/Final body weight) × 1000
of rats sera
Effect of the acetogenin-rich fraction of A. muricata leaves on
2.6. Biochemical analyses PSA levels of the rats with experimentally induced BPH is
shown in Figure 1. At the end of the 28 days of subcutaneous
The level of PSA in the rat sera was determined following the injection of testosterone propionate, there was a significant
Elabscience enzyme-linked immunosorbent assay (ELISA) test increase (p < 0.05) in PSA level (initial PSA) in groups that
kit procedure. The prostate protein content of the rats was received the hormone compared to the normal control, indi-
estimated by the method of Tiez [32]. Lipid peroxidation cating successful induction of BPH. Daily administration of the
was estimated by measuring thiobarbituric acid reactive sub- acetogenin fraction, at the dose level of 200 mg/kg body
stance expressed in terms of malondialdehyde (MDA) accord- weight to the hyperplasic rats for seven days, caused a signifi-
ing to the method described by Wallin et al. [33]. The serum cant (p < 0.05) decrease in PSA level (final PSA) compared to
antioxidant activities: superoxide dismutase (SOD), gluta- the untreated rats (TP group). This result was not significantly
thione peroxidase (GPx) and catalase (CAT) were assayed by different (p > 0.05) from those treated with the standard BPH
the methods of Arthur and Boyne [34], Paglia and Valentine drug (dutasteride). The ameliorating effect of the plant
[35] and Sinha [36], respectively. Reduced glutathione (GSH) material on the induced BPH was in a dose-dependent
was evaluated by the method of Exner et al. [37]. manner.

2.7. Histopathologic examination 3.2. Effect of the acetogenin-rich fraction on prostate

and relative prostate weights of the rats
The dorsolateral lobes of the prostates were fixed in 10%
phosphate formalin solution for 48 h. The prostate tissues A significant increase (p < 0.05) in prostate weight and relative
were then dehydrated in grades of ethanol, cleaned in prostate weight was observed in the group that received the
xylene and embedded in paraffin wax. Each block of the hormone alone when compared to the normal control, also
tissue was sectioned at 5 µm using rotary microtone and validating that prostate hyperplasia was induced. A significant
stained with hematoxylin and eosin (H and E). The sections decrease (p < 0.05) in prostate weight, as well as relative

Figure 1. Effect of acetogenin-rich fraction of Annona muricata leaves (AFAL) on prostate-specific antigen (PSA) levels of testosterone propionate (TP)-induced BPH
in rats. Values are expressed as mean ± standard error of mean (n = 5). #Significant when compared to normal control (p < 0.05); *significant compared to TP control
(p < 0.05).
 REDOX REPORT 83

prostate weight, was observed in the group treated with the basal cell (BC) and eosinophilic secretion (ES) within the
acetogenin fraction when compared to the untreated rats alveoli. In the group that received the hormone alone, there
(TP group). This indicated a remedial effect of the plant was a severe hyperplastic gland (HG), change in morphology
material on the induced BPH. Also, there was no significant of the BC and secretion atrophy within the lumen. Treatment
difference (p > 0.05) in prostate and relative prostate of the rats with 200 mg/kg A. muricata acetogenin fraction
weights between the test rats and those treated with the showed near-normal architecture with well-perfused
reference drug (TP + Dutasteride group) Figure 2. secretion. This restoration is similar to the histoarchitecture
of prostate in group treated with a standard drug, Figure 6.

3.3. Effect of the acetogenin-rich fraction on prostate

protein content
4. Discussion
The results showed that BPH induction significantly increased
(p < 0.05) the prostate protein content as observed in the Medicinal plants remain one of the potent sources of human
group that received the hormone alone, when compared to health due to the bioactive compounds that are responsible
normal control. A significant decrease in prostate protein for their various pharmacological activities. A. muricata is a tra-
content was observed in the group treated with 200 mg/kg ditional medicinal plant that has been reported to possess
of the A. muricata acetogenin fraction when compared to acetogenins as major phytoconstituents [38]. These chemical
the TP group. This value was not significantly different from compounds have been shown to exhibit various biological
the group treated with the standard drug. However, a non-sig- activities including the cytotoxic effect against the neoplastic
nificant decrease (p > 0.05) in the prostate protein content cells which suggested their potential use as an antitumor
was observed in the group that received a lower concen- agent [39].
tration of the fraction (100 mg/kg) when compared to the In this study, BPH induction by subcutaneous adminis-
TP group (Figure 2). tration of testosterone propionate (3 mg/kg) to the rats for
28 days was successful as evidenced in the rise of serum
PSA level and prostate weights of rats that received the
3.4. Effect of the acetogenin-rich fraction on oxidative hormone alone. PSA is a glycoprotein that is predominately
stress parameters of the rat sera formed in the prostate gland and a reliable marker for BPH
[40]. A significant reduction in the final PSA observed in test
The effect of A. muricata acetogenin fraction on malondialde-
rats treated with the acetogenin-rich fraction of A. muricata
hyde (MDA) concentration and the activities of antioxidant
enzymes were estimated as indices for oxidative stress
status of the rats. Subcutaneous injection of testosterone pro-
pionate significantly increased the lipid peroxidation marker
(MDA) of the group that received the hormone alone and as
well decreased their antioxidant activities compared to the
normal control group. Administration of the acetogenin frac-
tion (200 mg/kg) reversed the impact of the exogenous
hormone as the MDA concentration significantly reduced
(Figure 3) and the antioxidant activities of the rat were elev-
ated (Figures 4 and 5). The obtained values were comparable
to the normal control and group administered the standard
drug.

Figure 3. Effect of acetogenin-rich fraction of Annona muricata leaves (AFAL) on

3.5. Histologic examination malondialdehyde (MDA) concentration in testosterone propionate (TP)-induced
BPH in rats. Values are expressed as mean ± standard error of mean (n = 5). #Sig-
The histologic section of prostate in normal group showed a nificant when compared to normal control (p < 0.05); *significant compared to
standard prostatic architecture with prostate gland lined by TP control (p < 0.05).

Figure 2. Effect of acetogenin-rich fraction of Annona muricata leaves (AFAL) on prostate weight, relative prostate weight and prostate protein content in testos-
terone propionate (TP)-induced BPH in rats. Values are expressed as mean ± standard error of mean (n = 5). #Significant when compared to normal control (p < 0.05);
*significant compared to TP control (p < 0.05).
84 P. N. OGBU ET AL.

Figure 4. Effect of acetogenin-rich fraction of Annona muricata leaves (AFAL) on superoxide dismutase (SOD) and catalase (CAT) activities in testosterone propionate
(TP)-induced BPH in rats. Values are expressed as mean ± standard error of mean (n = 5). #Significant when compared to normal control (p < 0.05); *significant com-
pared to TP control (p < 0.05).

Figure 5. Effect of acetogenin-rich fraction of Annona muricata leaves (AFAL) on glutathione peroxidase (GPx) activity and reduced glutathione (GSH) concentration
in testosterone propionate (TP)-induced BPH in rats. Values are expressed as mean ± standard error of mean (n = 5). #Significant when compared to normal control
(p < 0.05); *significant compared to TP control (p < 0.05).

(200 mg/kg body weight) is a pointer to the ameliorative observation; the relative prostate weight of rats that received
effect of the plant material on the induced BPH. Decrease of hormone alone increased significantly in comparison with
PSA level is associated with a reduction of prostatic hyperpla- normal control, but treatment with the acetogenin fraction
sia as a direct consequence of 5α-reductase inhibition [41]. reduced the relative prostate weight to a near-normal weight.
Also, persisted high level of PSA (final PSA) in the group BPH has been associated with increase in prostate protein
that received the hormone alone without any treatment is content [43] and the observation of the present study was in
an indication that the observed drop in PSA level of the accordance; rats that were injected with the hormone alone
treated rats was not due to self-recovery. Comparing the had elevated prostate protein content. Administration of the
level of decrease in the final PSA after the 7 days treatment BPH rats with acetogenin fraction of A. muricata significantly
suggests that AFAL (200 mg/kg body weight) exhibited a decreased the protein content of their prostate which trans-
similar ameliorative effect as dutasteride on the induced lates to attenuation of the induced hyperplasia.
BPH, possibly through 5α-reductase inhibitory activity The histologic findings from this study also affirmed a
among other mechanisms. possible anti-BPH effect of the acetogenin-rich fraction of
Furthermore, prostate enlargement is also an important A. muricata, as prostatic histoarchitecture of the rats that
marker for BPH [42]. The enlargement is due to the increase received 200 mg/kg of AFAL showed recovery from prostatic
in both epithelial and stromal cell number (hyperplasia), hyperplasia when compared to the untreated rats.
resulting in an increase of prostate weight. Thus, the signifi- Several literatures have associated oxidative stress with
cant reduction of prostate weight of the BPH rat to near BPH development [23]; this could arise as a result of overpro-
normal after treatment with the acetogenin-rich fraction duction of oxidant molecules or depletion in the antioxidant
(200 mg/kg) is also suggestive of a remedial effect of the system during prostate enlargement. Besides the possible
plant material on the induced BPH. The results of the relative prostate tissue damage by the reactive oxygen species, a
prostate weights are equally in agreement with the above compensatory cellular proliferation can as well set in,
 REDOX REPORT 85

Figure 6. Micrograph of prostates in the experimental rats (Stain: H & E; Magnification: ×400). TP = testosterone propionate, AFAL = acetogenin-rich fraction of A.
muricata leaves, ES = eosinophilic secretion, BC = basal cell and HG = hyperplastic gland.

thereby worsening the prostate enlargement [23]. A balance however, are recommended to elucidate other possible
between oxidative stress and antioxidant system of the mechanisms of action of this plant material on BPH.
cells, however, plays an important role in the development
of prostate disease [44]. In this study, a significant elevation
5. Conclusion
in the concentration of MDA with a corresponding decrease
in the antioxidant activities observed in the rats that received The results of this study confirmed that acetogenin-rich frac-
the hormone alone confirmed associated oxidative stress in tion isolated from Annona muricata leaves could alleviate
BPH development. This could be due to the impact of the BPH induced in Wistar rats. At the dose of 200 mg/kg, the
supernormal dose of the exogenous hormone and physiologi- extract clearly reversed the effect of the supernormal dose
cal changes associated with prostate enlargement. However, of testosterone propionate on the Wistar rats. Enhanced anti-
administration of AFAL significantly decreased the MDA con- oxidant system may be one of the mechanisms through
centration and restored the antioxidant activities of the test which the plant material was able to exert its effect on the
rats in a dose-dependent manner. induced BPH. Therefore, acetogenin-rich fraction from
Dutasteride is a known potent BPH drug whose mechan- Annona muricata leaves may be useful in the management
ism of action is specific for the inhibition of 5α-reductase of BPH.
[45]. This was evident in this study where the drug obviously
exhibited a higher ameliorative effect in BPH markers (PSA
6. Ethics approval
levels, prostate weight, relative prostate weight and prostate
protein content) compared to AFAL, though the differences The laboratory animal protocol used in this study was
were not statistically significant. However, superior antioxi- approved by the Department of Biochemistry, University of
dant capacity of rats treated with AFAL (200 mg/kg dose) Nigeria, Nsukka as well as the Faculty of Biological Sciences
compared to the standard drug is notable. This suggests Ethics and Biosafety Committee of the same institution. The
that reinforcement of the antioxidant system could be one procedures agree with the guideline by the National Institutes
of the possible mechanisms through which the plant material of Health (NIH) publication on Guide for the Care and Use of
was able to attenuate the induced BPH. Further studies, Laboratory Animals.
86 P. N. OGBU ET AL.

Acknowledgements with natural products: Perspective of new pharmacological agents.

Inflamm Allergy Drug Targets. 2012;11:207–221.
This work did not receive any external funding. [20] Pagano E, Laudato M, Griffo M, et al. Phytotherapy of benign pro-
static hyperplasia. A minireview. Phytother Res. 2013;28(7):949–955.
[21] Dreikorn K. Phytotherapeutic agents in the treatment of benign pro-
Disclosure statement static hyperplasia. Curr Urol Rep. 2000;1:103–109.
[22] Gecit I, Meral I, Aslan M, et al. Peripheral mononuclear leukocyte
No potential conflict of interest was reported by the author(s). DNA damage, plasma prolidase activity, and oxidative status in
patients with benign prostatic hyperplasia. Redox Rep. 2015;20
(4):163–169.
ORCID [23] Minciullo PL, Inferrera A, Navarra M, et al. Oxidative stress in benign
prostatic hyperplasia: a systematic review. Urol Int. 2015;94:249–254.
Patience N. Ogbu http://orcid.org/0000-0001-6774-634X [24] Roduan MRM, Hamid RA, Kqueen CY, et al. Cytotoxicity, antitumor-
Ikechukwu M. Ogbu http://orcid.org/0000-0002-9053-7519 promoting and antioxidant activities of Annona muricata in vitro. J
Herb Med. 2019;15(2):100219.
[25] Baskar R, Rajeswari V, Kumar TS. In vitro antioxidant studies in leaves
of Annona species. IJEB. 2007;45:480–485.
References
[26] Mulia K, Winarcahyo SW, Krisanti E, et al. Practical isolation of Bullatacin
[1] Kapoor A. Benign prostatic hyperplasia management in the primary from Annona muricata leaves extract using an open column chromato-
care setting. Can J Urol. 2012;19(1):10–17. graphy Technique. Adv Mater Res. 2013;789:545–550.
[2] Kim JH, Park KM, Lee JA. Herbal medicine for benign prostatic hyper- [27] Pimenta LPS, Garcia GM, Gonçalves SGV, et al. In vivo antimalarial
plasia: A protocol for a systematic review of controlled trials. efficacy of acetogenins, alkaloids and flavonoids enriched fractions
Medicine (Baltimore). 2019;98(1):e14023. from Annona crassiflora Mart. Nat Prod Res. 2014;28(16):1254–1259.
[3] Jadallah S, Li Y, Chu LW, et al. Prevalence of BPH and lower urinary [28] Yang C, Gundala SR, Mukkavilli R, et al. Synergistic interactions
tract symptoms in West Africans. Prost Cancer Prost Dis. among flavonoids and acetogenins in Graviola (Annona muricata)
2012;15:170–176. leaves confer protection against prostate cancer. Carcinogenesis.
[4] Briganti A, Umberto C, Nazareno S, et al. Benign prostatic hyperpla- 2015;36(6):656–665.
sia and its aetiologies. Eur Urol Suppl. 2009;8:865–871. [29] Mulia K, Krisanti E, Maulana T, et al. Selective polarity-guided extrac-
[5] Ejike CECC, Eze KC. Prevalence of symptoms of benign prostatic tion and purification of acetogenins in Annona muricata L. leaves. Int
hyperplasia in Umudike and its relationship with measures of J Technol. 2015;7:1221–1227.
obesity. Asia Pac J Clin Nutr. 2015;7(1):1–8. [30] Lorke D. A new approach to practical acute toxicity testing. Arch
[6] Kaplan SA, Roehrborn CG, Rovner ES. Tolterodine and tamulsolin for Toxicol. 1983;53:275–289.
treatment of men with lower urinary tract symptoms and overactive [31] Shin IS, Lee MY, Ha HK, et al. Inhibitory effect of Yukmijihwang-tang,
bladder: a randomized controlled trial. JAMA. 2006;296:2319–2328. a traditional herbal formula against testosterone induced benign
[7] Nahata A, Dixit KV. Sphaeranthus indicus attenuates testosterone prostatic hyperplasia in rats. BMC Compl Alternative Med.
induced prostatic hypertrophy in albino rats. Phytother Res. 2012;12:48.
2011;25:1839–1848. [32] Tietz NW. Clinical Guide to laboratory tests. Philadelphia: W.B.
[8] Geavlete P, Multescu R, Geavlete B. Serenoa repens extract in the Saunders, Co.; 1995. p. 216–580.
treatment of benign prostatic hyperplasia. Ther Adv Urol. [33] Wallin B, Rosengren B, Shertzer HG, et al. Lipoprotein oxidation and
2011;3:193–198. measurement of thiobarbituric acid reacting substances (TBARS) for-
[9] Heidenreich A, Bellmunt J, Bolla M, et al. Guidelines on prostate mation in a single microtitre plate: Its use for evaluation of antioxi-
cancer. part 1: screening, diagnosis, treatment of clinically localized dants. Anal Biochem. 1993;208:10–15.
disease. Eur Urol. 2011;59:61–71. [34] Arthur JR, Boyne R. Superoxide dismutase and glutathione peroxi-
[10] Kalu WO, Okafor PN, Ijeh II, et al. Effect of kolaviron, a biflavanoid dase activities in neutrophils from selenium deficient and copper
complex from Garcinia kola on some biochemical parameters in deficient cattle. Life Sci. 1985;36:1569–1575.
experimentally induced benign prostatic hyperplasic rats. Biomed [35] Paglia DE, Valentine WN. Studies on the quantitative and qualitative
Pharmacother. 2016;83:1436–1443. characterization of erythrocyte glutathione peroxidase. J Lab Clin
[11] Sharma M, Chadha R, Dhingra N. Phytotherapeutic agents for benign Med. 1967;70:158–169.
prostatic hyperplasia: an overview. Mini Rev Med Chem. 2017;17 [36] Sinha AK. Colorimetric assay of catalase. Anal Biochem. 1972;47:389–
(14):1346–1363. 394.
[12] Fujita R, Liu J, Shimizu K, et al. Anti-androgenic activities of [37] Exner R, Wessner B, Manhart N, et al. Therapeutic potential of gluta-
Ganoderma lucidum. J Ethnopharmacol. 2005;102:107–112. thione. Wien Klin Wochenschr. 2000;112:610–616.
[13] Xu Y, Ventura S. Extracts of bark from the traditional Chinese herb [38] Padma P, Chansouria J, Khosa R. Effect of alcohol extract of Annona
Phellodendron amurense inhibits contractility of the isolated rat pros- muricata on cold immobilization stress induced tissue lipid peroxi-
tate gland. J Ethnopharmacol. 2010;127:196–199. dation. Phytother Res. 1997;11:326–327.
[14] Levy A, Sivanesan D, Murugan R, et al. Urtica dioica Induces cytotox- [39] Gajalakshmi S, Vijayalakshmi S, Rajeswari DV. Phytochemical
icity in human prostate carcinoma cells: involvement of oxidative pharmacological properties of Annona muricata: a review. Int J
stress. Mitochondrial depolarization and apoptosis. Trop J Pharm Pharm Pharm Sci. 2012;4(2):0975–1491.
Res. 2014;13(5):711–717. [40] Choi H, Jung Y, Park J, et al. Cinnamomi cortex (Cinnamomum verum)
[15] Coria-Tellez AV, Montalvo-Go E, Yahia EM, et al. Annona muricata: a suppresses testosterone-induced benign prostatic hyperplasia by
comprehensive review on its traditional medicinal uses, phytochem- regulating 5α-reductase. Sci Rep. 2016;6:31906.
icals, pharmacological activities, mechanisms of action and toxicity. [41] Sing B, Ram SN, Pandey VB, et al. Studies on antiinflammatory
Arab J Chem. 2018;11:662–691. activity of taraxasterol acetate from Echinops echinatus in rats and
[16] Adewole SO, Caxton-Martins EA. Morphological changes and hypo- mice. Phytother Res. 1991;5:103–106.
glycemic effects of Annona muricata Linn. (Annonaceae) leaf [42] Shin I, Lee M, Jung D, et al. Ursolic acid reduces prostate size and
aqueous extract on pancreatic B-cells of streptozotocin-treated dia- dihydrotestosterone level in a rat model of benign prostatic hyper-
betic rats. Afr J Biomed Res. 2006;9:173–187. plasia. Food Chem Toxicol. 2012;50:884–888.
[17] Moghadamtousi SZ, Fadaeinasab M, Nikzad S, et al. Annona muricata [43] Ejike CECC, Ezeanyika LUS. Inhibition of the experimental induction
(Annonaceae): a review of its traditional uses, isolated acetogenins of benign prostatic hyperplasia: a possible role for fluted pumpkin
and biological activities. Int J Mol Sci. 2015;16(7):15625–15658. (Telfairia occidentalis Hook f.) seeds. Urol Int. 2011;87:218–224.
[18] Asare GA, Afriyie D, Ngala RA, et al. Antiproliferative activity of [44] Khandrika L, Kumar B, Koul S, et al. Oxidative stress in prostate
aqueous leaf extract of Annona muricata L. on the prostate, BPH-1 cancer. Cancer Lett. 2009;282:125–136.
cells, and some target genes. Integr Cancer Ther. 2015;14(1):65–74. [45] Djavan B, Milani S, Fong YK. Dutasteride: a novel dual inhibitor of 5α-
[19] Azimi H, Khakshur A-A, Aghdasi I, et al. A review of animal and reductase for benign prostatic hyperplasia. Expert Opin Pharmacol.
human studies for management of benign prostatic hyperplasia 2005;6(2):311–317.