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Chinese Medicine, 2014, 5, 1-6
Published Online March 2014 in SciRes. http://www.scirp.org/journal/cm
http://dx.doi.org/10.4236/cm.2014.51001
How to cite this paper: Jahromy, M.H., Khakpour, S. and Khorgami, Z. (2014) The Antidepressant-Like Effects of Punica
granatum (Pomegranate) Extract in Mice. Chinese Medicine, 5, 1-6.
http://dx.doi.org/10.4236/cm.2014.51001
The Antidepressant-Like Effects of Punica
granatum (Pomegranate) Extract in Mice
Mahsa Hadipour Jahromy1*, Shahrzad Khakpour2, Zhinoos Khorgami3
1Department of Pharmacology, Tehran Medical Sciences Branch, Islamic Azad University, Tehran, Iran
2Department of Physiology, Tehran Medical Sciences Branch, Islamic Azad University, Tehran, Iran
3Iranian Tissue Bank and Research Center, Tehran University of Medical Sciences, Tehran, Iran
Email: *Jahromymh@yahoo.com
Received 9 December 2013; revised 18 January 2014; accepted 3 February 2014
Copyright © 2014 by authors and Scientific Research Publishing Inc.
This work is licensed under the Creative Commons Attribution International License (CC BY).
http://creativecommons.org/licenses/by/4.0/
Abstract
The aim of the present work is to evaluate the putative antidepressant-like effects of pomegranate
fruit extract including seeds (PFE) on the performance of male mice in the forced swimming test
(FST), after acute administration, after short-term treatment (7 days) and, after repeated admin-
istration in a 24-h period (24, 12 and 1 h before swimming test). A single dose (20 ml/kg p.o.) of
PFE, in male mice provoked a significant reduction of the immobility time. Such effect was also
observed with short-term treatment (7 days) with doses of 1 and 10 ml/kg/day of PFE. Moreover,
it was noted that there were important differences in the onset of the antidepressant-like effect in
the FST, depending on the modality of treatment with PFE. Both efficacy and potency were higher
when repeated administration of PFE was used, and surprisingly the dose of 10 ml/kg (24, 12 and
1 h before swimming test) was as effective as Fluoxetine. In the same way, the short term admin-
istration (7 days) improved significantly efficacy and potency of the PFE in comparison to a single
dose treatment. These results indicate an antidepressant-like profile of action for PFE which de-
serves further research.
Keywords
Pomegranate; Antidepressant Effect; Forced Swimming Test
1. Introduction
Depression is a major disease affecting nearly 13% - 20% of the population [1]. Roughly, 90% of patients with
depressive symptoms suffer from mild to moderate depression, while only 10% are severely depressed. In mild
*
Corresponding author.
M. H. Jahromy et al.
2
to moderate depression, in particular, some practitioners and patients are reluctant to use standard antidepres-
sants like tricyclic antidepressants or specific serotonin reuptake inhibitors (SSRIs), because of their side effects.
Therefore, additional treatment strategies with fewer side effect profile, credible benefits and moderate costs are
of particular interest [2] [3]. In the search for new molecules useful for the treatment of neurological disorders,
medicinal plant research, worldwide, has progressed constantly, demonstrating the pharmacological effective-
ness of different plant species in a variety of animal models [4]. At the present time, forced swimming test (FST)
is the most widely used model for assessing potential antidepressant activity in rodents, following acute and
short-term treatment [5].
Pomegranate (Punica granatum), a small tree originating in the Orient, belongs to the Punicaceae family [6].
It is grown mainly in Iran, India and in most Near and Far East countries. The main use of pomegranate is as ta-
ble fruit, but large amounts are used in the beverage and liquor industries [7]. In folk medicine, pomegranate
preparations have a number of therapeutic actions [8].
The edible part of pomegranate is rich in anthocyanins thus; the major class of phytochemical present in po-
megranate is the polyphenols and includes flavonoids, condensed tannins and hydrolysable tannins. Hydrolysa-
ble tannins are predominant polyphenols found in pomegranate juice and account for 92% of its antioxidant ac-
tivity [9] [10].
The present research was undertaken to study the duration of immobility in mice in the FST, after acute ad-
ministration, after short-term treatment (7 days) and, after repeated administration in a 24-h period (24, 12 and 1 h
before swimming test) following oral ingestion of a standardized preparation of pomegranate fruit extract (PFE).
2. Materials and Methods
2.1. Pomegranate Extract
Pomegranates (Punica granatum L.), was collected by one of the colleagues from the agriculture garden (under
supervision of Agricultural Research Organization in Fars Province) in Shiraz, a city where known to have one
of the best pomegranate native in Iran. Then washed, chilled to 4˚C, and stored. The seeds of the fruit containing
the intact juice sacs were manually separated from the pericarp, and the whole juice extracted by the aid of elec-
tric juicer so that seeds break. Then filtered and stored in clean jars in fridge.
2.2. Animals
Male BALB/c mice, weighing 25 - 30 g were used in these experiments. They were group housed under the fol-
lowing laboratory conditions: temperature 23˚C ± 1˚C, humidity 40% - 60%, 12 h:12 h L/D cycle, lights on at
07:00 h. Food and water were available ad libitum. All the experiments were carried out between 10:00 and
15:00 h in testing rooms adjacent to the animal rooms. Each experimental group consisted of ten mice. Mice
were treated in accordance with the current law and the NIH Guide for the Care and Use of Laboratory.
2.3. Drugs
Fluoxetine (Sigma) were suspended in normal saline (0.9% NaCl).
2.4. Behavioral Tests
This test was performed according to the procedure described by Porsolt et al. [11]-[13], with slight modifica-
tions [14]. Briefly, 1 h after dosing, the animals were individually forced to swim in a transparent glass vessel
(25 cm high, 15 cm in diameter) filled with (12.5 cm high) water at 21˚C - 24˚C. The total duration of immobil-
ity (in seconds) was measured during the last 4 min of a single 6 min test session. Five groups of 10 mice were
treated acutely with single dose of vehicle, PFE (1, 10 and 20 ml/kg) p.o. (Gavage), Fluoxetine i.p. (10 mg/kg)
and 1 h later mice were individually forced to swim in the glass vessel. Other five groups of mice were short-
term treated (7 days) with a single daily dose of vehicle, PFE (1, 10 and 20 ml/kg) p.o., or Fluoxetine i.p. (10
mg/kg), and 1 h after the last administration they were individually forced to swim in same conditions. In a dif-
ferent set of experiments, five groups of 10 animals were subjected to repeated administration of three doses of
vehicle, PFE using 1, 10 and 20 ml/kg p.o., Fluoxetine i.p. (10 mg/kg), 24, 12 and 1 h prior to the swimming test.
Behaviour was monitored from the frontal side by a video camera for subsequent analysis. Mice were consid-
M. H. Jahromy et al.
3
ered immobile when they made no further attempts to escape except the movements necessary to keep their
heads above the water.
2.5. Statistical Analysis
The results are expressed as mean ± S.D, and statistical analysis of the data was performed by Origin VI. ***p <
0.001; **p < 0.01 were considered significantly different from vehicle, after non-parametric ANOVA.
3. Results
Effects of PFE on the Immobility Time
A significant shortening of the immobility time was observed when an acute dose of either 20 ml/kg p.o., of PFE
(p < 0.01) or fluoxetine (10 mg/kg i.p.) (p < 0.001) was administered to mice compared to vehicle treated ani-
mals. The reduction of immobility time, between these two mentioned groups was at the same level, statistically
(Figure 1). Short-term treatment (7 days) induced a decrease of the immobility time when the animals were ex-
posed to the FST after dosing orally, once a day, with 1 (p < 0.001) and 10 (p < 0.001) ml/kg of PFE (Figure 1).
In addition, repeated administration in a 24 h period showed a significant reduction of the immobility time with
1 (p < 0.01) and 10 ml/kg (p < 0.001) of PFE in different groups of mice submitted to FST in comparison to
control group (Figure 2).
4. Discussion
The present study characterized the effects of the administration of PFE on mice performance in FST following
acute, short-term and repeated treatment. In many previous papers the antioxidant and anti-inflammatory prop-
erties of pomegranate fruit juice in animal labs has been emphasized [15]-[18]. Also, it has been reported that
Pomegranate extract improves a depressive state and bone properties in menopausal syndrome model ovariec-
tomized mice [19]. Besides that, it was demonstrated that PFE exhibits low toxicity, no lethality, was well toler-
ated, did not induce significant changes in several behavioural and physiological parameters and is devoid of any
hypnosedative activity, when administered to mice [15]. Recently, the antidepressant action of some herbal medi-
cines has been reported focusing on polyphenols as their major component [3] [4] [14]. The edible part of pome-
granate is rich in anthocyanins thus; the major class of phytochemical present in pomegranate is the polyphenols.
Figure 1. Effect of the vehicle, fluoxetine (10 mg/kg, i.p.) and increasing single
doses of PFE (1, 10 and 20 ml/kg p.o) on the forced swimming test (FTS) in mice.
Each bar represents the mean ± SD of 10 animals. ***p < 0.001; **p < 0.01, signifi-
cantly different from vehicle, after non-parametric ANOVA.
01234
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50
100
150
200
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*
*
*****
*
*
**
*
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Immobility Time (sec)
Vehicle, Flouxetine, PFE 1 ml/kg, 10 ml/kg, 20 ml/kg
One Day Treatment
Seven Days Treatment
M. H. Jahromy et al.
4
Figure 2. Effect of repeated administration of three doses in 24 h of vehicle, fluoxetine
(10 mg/kg i.p) and increasing doses of PFE (1, 10, and 20 ml/kg p.o) on the forced
swimming test (FTS) in mice. Each bar represents the mean ± SD of 10 animals. ***p <
0.001; **p < 0.01, significantly different from vehicle, after non-parametric ANOVA.
Because the pharmacotherapy of depression typically requires chronic drug treatment to obtain a full response
in terms of antidepressant effect, it is critical to perform, not only acute, but also short-term, chronic and re-
peated treatments in the FST mice model. Certainly, the administration of PFE produced a diminution of immo-
bility time of mice exposed to the FST under acute, short-term or repeated modalities. Indeed, single dose of 20
ml/kg p.o., of PFE provoked a highly significant reduction of immobility time (p < 0.01). Similar results were
also observed with short-term treatment (7 days) with single dose of 1 (p < 0.001), and 10 (p < 0.001) ml/kg/day
of PFE, but not with higher dose. Additionally, in a different set of experiments, repeated administration in a
24-h period (24, 12 and 1 h before swimming) doses of 1 (p < 0.01) and 10 (p < 0.001) ml/kg p.o., of PFE pro-
voke significant reduction of the immobility time of male mice subjected to the FST, when compared to control
group (vehicle) but not with higher dose. It is important to note that although single acute high dose, showed an-
tidepressant effect as fluoxetine, no significant effects observed in high dose of 20 mg/kg of PFE in repeated
administration. This can be interpreted as acute tolerance or tacyfilaxy that might have happened in receptor re-
sponses. Moreover, depending on the modality of treatment with PFE (acute, short-term or repeated) in the FST,
it was observed important differences in the onset of the antidepressant-like effect. In fact, both efficacy and po-
tency of PFE as antidepressant were higher when repeated administration was used, and interestingly 10 mg/kg
(24, 12 and 1 h before swimming) was as effective as fluoxetine (10 mg kg i.p.) used as positive control. In the
same way, the short term administration (7 days), improves significantly the efficacy and potency of the PFE, in
comparison to single dose treatments. These behavioural effects were similar to those seen in mice treated with
conventional antidepressant drugs, such as tricyclic, monoamine oxidase inhibitors and selective serotonine re-
uptake inhibitors agents [5] [11] [20].
5. Conclusion
In conclusion, the results of this preclinical study provide evidence about the antidepressant effects of the po-
megranate fruit extract administered either acute or repeatedly in mice. These results contribute to the scientific
validation of the indications of this plant in Iranian folk medicine. However, further experiments are needed to
identify its active compounds and the corresponding mechanisms of action. Our results encourage us to pursue
the identification of the molecules associated to the effect observed in PFE.
Acknowledgements
The authors would like to thank M. Aghakhani, for his help during experiments. The experimental work has
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Immobility Time (sec)
Vehicle Fluoxetine PFE 1 ml/kg PFE 10 ml/kg PFE 20 ml/kg
M. H. Jahromy et al.
5
been done at physiology and pharmacology laboratory of medical sciences research center, Tehran Medical
Sciences Branch, Islamic Azad University.
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Note List of Abbreviations
Pomegranate fruit extract (PFE)
Forced swimming test (FST)
Specific serotonin reuptake inhibitors (SSRIs)
