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Advances in Environmental Biology, 7(14) December 2013, Pages: 4905-4908
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Advances in Environmental Biology
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Investigating Effects of Alcohol Aqueous Extract of Harmala Seed on the Plasma Levels
of Pituitary - Thyroid Hormones Axis and Liver Enzymes and Body Weight in Adult
Male Rats
1,2
Amineh Daneshi, 2,3Kargar Jahromi Hossein, 4Ghorishi Fakhrossadat, 4Zahedi Ali, 5Bathaee Seyed
Hamid, 6Saberi Roza, 7Farzam Mohammad
1
The Departmant General of Yasuj Province Education.
Zoonoses research center, Jahrom University of Medical Sciences, Jahrom, Iran.
3
Young Researchers Club Elite, Jahrom Branch, Islamic Azad University, Jahrom, Iran.
4
The Departmant General of Bandar Abbas Province Education.
5
Institution of Supreme Education and Industry of Maragheh, Iran.
6
Department of Biology, Islamic Azad University, Arsanjan, Fars, Iran.
7
Department of Anatomy and Embryology, International Branch, Shiraz University, Shiraz, Iran.
2
ARTICLE INFO
Article history:
Received 22 October 2013
Received in revised form 14
January 2014
Accepted 20 January 2014
Available online 15 February 2014
Key words:
Thyroid Stimulating Hormone,
Thyroxine,Triiodothyronine,
Harmala.
ABSTRACT
Introduction and Objectives: Harmala plant has activations of anti-bacterial, anti-fungal
and anti-tumor, and inhibit Monoamine oxidase enzyme, stimulate nervous system and
modulate activation of Endocrine. The aim of this study was to investigate the effect of
harmala seed extract on the levels of hormones of pituitary-thyroid and liver enzymes.
Materials and methods: This research is an experimental study that was done on 50
adult male Wistar rats. In this study, animals were divided into 5 groups. The control
group did not receive any treatments, the sham group for 14 consecutive days received
orally 1 ml of distilled water and experimental groups for 14 consecutive days received
respectively 90 mg/kg, 180 mg/kg and 270 mg/kg harmala seed extract that was given
orally and on the fifteenth day, stimulating hormones concentration of thyroid,
thyroxine and triiodothyronine and liver enzymes were measured and the results
analyzed using SPSS software and ANOVA test. Results: Harmala seed extract at the
minimum and maximum doses significantly reduced the rate of thyroid stimulating
hormone, and in medium and maximum doses significantly reduced the rate of
thyroxine and triiodothyronine, but body weight and liver enzymes did not show
significant changes (P<0.05). Conclusion: The results indicate that the harmala seed
extract can be used in the treatment of hyperthyroidism.
© 2013 AENSI Publisher All rights reserved.
To Cite This Article: Amineh Daneshi, Kargar Jahromi Hossein, Ghorishi Fakhrossadat, Zahedi Ali, Bathaee Seyed Hamid, Saberi Roza,
Farzam Mohammad., Investigating Effects of Alcohol Aqueous Extract of Harmala Seed on the Plasma Levels of Pituitary - Thyroid
Hormones Axis and Liver Enzymes and Body Weight in Adult Male Rats. Adv. Environ. Biol., 7(14), 4905-4908, 2013
INTRODUCTION
Maintaining homeostasis of the body is the key point in human health that is controlled by several factors,
especially pituitary–thyroid hormone axis. Underactive and overactive thyroid with disruption in body health
imposes a high cost for patient, and in many cases there is no possible treatment [1]. From the past to now, in
medical and treatment of diseases medicinal plants have generally been used and peganum is one of the
medicinal plants with the scientific name of Peganum harmala, from the family of zigu filaseh. Peganumhermla
is a perennial grass that grows to a height of approximately 30 to 40 cm. This plant grows in semi-arid and arid
lands and has general distribution in Europe, Turkey, Iran, Afghanistan, Pakistan, India, Iraq, Syria, Jordan,
Lebanon, Palestine, Arabia and North Africa. It can be found in Iran in areas where there is little salt. This plant
is also found in America and Australia. Harmala seeds contain compounds such as Alkaloids like Harmaline,
Harmaine, Harmalole, Harman, Vasicine and Vasizinone [1]. Harmala traditionally produce effects such as
sleepiness, sweating, anti-parasite, binding rule, the embryo abortion, anti-cancer, anti-bacterial, nervous system
stimulants and inhibitors of the enzyme monoamine oxidase [2,3]. Harmala alkaloid composition of the seeds
of the neurotransmitters dopamine and serotonin stimulates a dose-dependent manner [4,5]. The most important
factor controlling the synthesis and secretion of thyroid hormones is applied by regulating hypothalamus–
pituitary hormonal axis. Thyroid-stimulating hormone, which is released from hypothalamus has an impact on
Corresponding Author: Hossein Kargar Jahromi, Jahrom University of Medical Sciences, Jahrom, Iran.
Tel: +989361554955
E-mail: [email protected]
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Hossein Kargar Jahromi et al, 2013
Advances in Environmental Biology, 7(14) December 2013, Pages: 4905-4908
anterior pituitary and causes secretion of thyroid stimulating hormone [6,7]. Thyroid hormones T4 (thyroxine)
and T3 (triiodothyronine) also affect the secretion of thyroid stimulating hormones that are secreted by adno
pituitary [8,9]. Thyroid hormones stimulate the transcription of numerous genes, intensify production of large
amounts of enzymes, structural proteins and carrier and increase the basal metabolic [10,11]. Because of the
many uses of this plant in traditional medicine and the effect of this plant in endocrine researches, other axes
have been considered but a study of the effect of harmala seed on performance of pituitary-thyroid hormone axis
associated with changes in liver enzymes and body weight has not been done, so the aim of the present study is
to investigate the influence of hydro-alcoholic seed extract on plasma levels of hormones pituitary-thyroid axis.
MATERIALS AND METHODS
In this study 50 adult male Wistar rats weighing approximately 260-320 gr were used. The samples were
divided into five groups of 10 that include control and sham groups and 3 experimental groups. Each of the
groups were in separate cages and were given one week to adapt to the laboratory environment. During the
prescribed period, all animals were given equal and unrestricted food and water at a constant temperature (25 +2
° C) and natural light (12 hours light, 12 hours dark). In the entire project all ethics in relation to working with
laboratory animals have been considered. In the spring season the required amount of harmala seeds was
collected from Nurabad-Mamasani in Iran. Then a perfect sample of the above-mentioned plant in order to
identify plant species in the herbarium of the Research Center of the Jahad university of Kohkiluyeh and Boyer
Ahmad province in Iran, Herbarium No. 4633 inside the country was recorded and coded. 500 gr of powdered
seeds were soaked in 96% alcohol and after 48 hours exposure on shaker machine, at room temperature and
away from heat and light the mixture was smoothed. The filtered solution was passed through Whatman filter
paper (number 1) for better refining. The resulting solution was used as raw material for the extraction stage and
was concentrated in a rotary machine in vacuum condition. The resulting material was placed in an oven for 5
days at 40 to 45°C. The final material weight of the above process was 94 gr. The animals were randomly
grouped. Animals in the control group did not receive any special treatment. The sham group received daily 1 cc
distilled water for 14 days orally and three experimental groups, 1-3 respectively received 90 mg/kg, 180 mg/kg
and 270 mg/kg of harmala seed extract orally. On day 15, after weighing, the animals were anesthetized by ether
and blood samples were taken directly from the heart. To obtain plasma, blood samples were taken into tubes
containing EDTA anticoagulant and then the required centrifuged blood plasma samples were kept frozen in the
freezer at - 20°C. Plasma samples for measuring stimulating hormones of thyroid, thyroxine and
triiodothyronine were assessed by radioimmunoassay method(12). Liver enzymes were measured by
spectrophotometric method. Data were analyzed by SPSS software and ANOVA test.
Results:
The results indicate that plasma levels of thyroid stimulating hormone in the receiving groups of minimal
doses (90 mg /kg) and maximum (270 mg/ kg) compared to control and sham groups had a significant decrease
(P≤0.05) while in the receiving dose of (180 mg/kg) groups there was no significant difference (Table 1). Also,
after measuring plasma levels of thyroxine and triiodothyronine hormones it was determined that the rate of
the above hormones, just in the groups receiving the intermediate (180 mg/kg) and maximum (270 mg/kg)
doses compared with control and sham groups showed significant decrease (P ≤0.05) (Table 1). These findings
suggest that increasing the rate of receiving extract has decreased the concentration of thyroxine hormone.
Scheffe test between experimental groups receiving medium and maximum dose, and control and sham groups
showed significant difference, but in other cases there was no significant difference (P≤0.05). Also, by
increasing the concentration of harmala extract the amount of triiodthyronine hormone in the rats under study
has decreased. Scheffe test between experimental groups receiving the intermediate and maximum dose, and
control and sham groups has shown significant decrease (P≤005), but between experimental groups 1-3 there
was no significant difference. Body weight (Table 2) and liver enzymes have not shown significant difference in
P≤0.05 levels in any of the groups with control group.
Table 1: Comparison of changes in plasma of pituitary-thyroid hormone axis in examined groups based on the mean and standard deviation
Variable
Thyroid stimulating hormone
Thyroxine hormone
Triiodothyronine
groups
(DI)
(DI)
hormone (DI)
Control
0.16+0.09
3.02+1.1
1.03+0.25
Sham
0.17+0.09
3.38+0.79
0.83+0.15
Experimental 1 (mg/kg)
0.13+0.04
2.56+0.69
0.71+0.08
Experimental 2 (180 mg/kg)
0.18+0.05
2.06+0.47
0.63+0.11
Experimental 3 (270 mg/kg)
0.12+0.04
1.78+0.64
0.62+0.11
The asterisk sign *indicates significant difference with control and sham groups
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Table 2: Changes in rats weight to the harmala extract received on the first, tenth and fifteenth days based on mean and standard deviation
Weight of fifteenth day
Weight of tenth day
Weight of first day
Concentration
group
315±47.464
300±49.752
294.4±47.879
Control
303.1±31.405
297.5±27.837
295.3±29.154
Sham
298.8±44.374
264.8±41.483
278.96±44.187
Experimental 1 (90 mg/kg)
284.7±36.174
282.2±32.117
273.2±31.371
Experimental 2 (180 mg/kg)
284.8±37.424
279.1±29.414
269.5±27.89
Experimental 3 (270 mg/kg)
Table 3: Changes in liver enzymes of the groups of receiving harmala extract.
AST(mean±SD)
ALT(mean±SD)
185.2±65.072
154.1±38.526
153.4±27.399
159.6±26.496
202.9±69.589
89±18.024
83.7±7.746
85.50±11.266
96.6±33.635
93.6±14.206
Concentration
group
control
sham
Experimental 1
Experimental 2
Experimental 3
Discussion and conclusion:
Harmala seed, having alkaloid compounds such as harmala, harmaline, harmalole, and steroidal compounds
of lygan and Saponins have many medicinal effects including regulation of endocrine functions [1]. Thyroid
disorders, if not treated properly impose irreparable damage to patients and in medicine a variety of methods
are used to treat thyroid disorders. The aim of this study was to investigate the effect of hydroalcoholic extract
of harmala seeds on plasma levels of the pituitary–thyroid hormone axis. The results showed harmala seed
extract has a lowering effect on hormone axis function of pituitary-thyroid. Secretion of thyroid stimulating
hormone is stimulated by thyrotropin-releasing hormone of Paraventricolarhypothalamus core. Several
neurotransmitter structures control secreting neurons of thyrotropin-releasing hormone in hypothalamus. One of
these structures of neurotransmitter, is neuropeptide Y [13]. Since the fibers in harmala seed extract through the
increase of leptin secretion have an inhibitory effect on activation of secreting neurons of neuropeptide Y, by
reducing the activity of neuropeptide-secreting neurons that have a stimulating effect on secretion of the
thyrotropin-releasing hormone, the rate of this neuro-hormone and so secretion of thyroid stimulating hormone
would decrease [14,15]. Regarding the stimulatory role of thyroid stimulating hormone on secretion of thyroid
hormones of thyroxine and triiodothyronine, plasma levels of thyroid hormones would decrease. Several studies
have shown that harmala is one of the alkaloids in the harmala seed extract that has inhibitory properties for
Monoamine oxidase enzyme and with regard to the inhibition of this enzyme, catecholamine neurotransmitters
levels (dopamine, norepinephrine, epinephrine) and Indole amine (serotonin) will increase. Further, since
serotonin is known as an inhibitory neurotransmitter in the secretion of thyrotropin-releasing hormone, harmala
seed extract, by increasing the rate of serotonin causes a decrease of secretion of thyrotropin-releasing hormone
and subsequently thyroid stimulating hormone levels in plasma, so thyroxine and triiodothyronine would
decrease [16]. Other studies have shown that dopamine, in the level of hypothalamus with decreased secretion
of thyrotropin-releasing hormone, and in the level of pituitary directly prevent secretion of thyroid stimulating
hormone, and causes decreased plasma levels of thyroid stimulating hormone, either free or bound to plasma
proteins. Harmala seed extract, also with inhibition of the enzyme monoamine oxidase, causes increase in the
rate of dopamine that can decrease plasma levels of pituitary-thyroid hormones axis [17]. Dopamine through
receptors can prevent secretion of thyroid stimulating hormone from the anterior pituitary. Also, dopamine by
stimulating somatostatin secretion can reduce thyroid stimulating hormone secretion, and so plasma levels of
hormones thyroxine and triiodothyronine would decrease [18]. Based on the investigation, steroid compounds
cause a decrease of thyroid hormones transporter proteins in serum [19]. Thus, despite the steroids in harmala
seed extract decrease of thyroxine and triiodothyronine hormones through decrease of thyroid hormones
transporter proteins is justified (20).According to the results in Table 2 it was determined that harmala does not
apply significant changes to body weight. Based on research conducted by Sebastian et al it was determined
that the combinations in harmala seed extract cause an increase in leptin in adult mammals and finally, by
inhibiting the secretion of NPY causes reduction of food intake, which is consistent with this research study
[15]. According to the results in Table 3, plasma concentration of enzymes ALT and AST in the control and
experimental and sham groups did not show significant changes. Regarding the fact that in the hypothyroidism
conditions through the damages that are incurred on the liver tissue, plasma levels of liver enzymes such as
ALT and AST would increase and Harmala seed extract reduces plasma levels of thyroid hormones, so the lack
of significant changes in plasma levels of these enzymes is justified [21]. According to the results of this study
the effect of harmala seed extract on pituitary-thyroid axis is examined, it is recommended that the combined
effect of harmala extract on this axis also be examined, as the findings indicate that the inhibitory effect of
compounds in harmala seed extract effect the secretion of thyroid hormone, so its use in overactive thyroid is
recommended.
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Hossein Kargar Jahromi et al, 2013
Advances in Environmental Biology, 7(14) December 2013, Pages: 4905-4908
ACKNOWLEDGEMENTS
The authors would like to express their appreciation to the staff of the Department of Anatomy of Medical
Science University of the Jahrom branch for their unwavering cooperation.
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