Central Nervous System
Depressant Activity of Aqueous
Extract of Leaves of Azadirachta indica Linn
in Mice
P. Thamarai Selvi*,
M. Senthil Kumar,
T. Yaswanth, E. Adiyaman,
P.T. Anusha
Department
of Pharmacology, Annai Veilankanni’s
Pharmacy College, Chennai-15, India
*Corresponding Author E-mail: tthamarai_pharma@yahoo.com.
ABSTRACT:
Aim of the study to investigate the effects of
Acetone extract of Azadirachta indica on central nervous system and possibility to use
it as folk medicine. The Acetone extract of Azadirachta
indica was soxhlat
extracted with Acetone and concentrated. The effect of Acetone extract on CNS
was studied by using analgesic activity and sedative-hypnotic activity.
Preliminary phytochemical evaluation of extract was
also carried out. The extract (100mg
and 200mg/kg, i.p) showed significant (p<0.001) potentiation of reaction time to thermal stimulus. It also
showed significant (p<0.01). Potentiation o phenobarbitone induced sleeping time to control.
KEYWORDS: Azadirachta indica, Analgesic activity, Sedative-hypnosis.
INTRODUCTION:
Currently available antipsychotics are associated1
with variety of autonomic, endocrine, allergic, haematopoietic,
and neurological side effects. As a result there is a high prevalence of usage
of complementary and alternative medicines for treatment of psychiatric
disorders. In the search for new therapeutic products for the treatment of neurological
disorders. Medicinal plant research worldwide has progressed constantly,
demonstrating the pharmacological effectiveness of different plant species in a
variety of animal models. Azadirachta indica
Linn (Meliaceae) commonly known as neem is a large tree upto 8m high
with almost straight trunk, distributed throughout the India2,
Pakistan, Bangladesh and Srilanka. The medicinal utilities have been described,
especially for leaf, fruit and bark. It is used for the treatment of
rheumatism, chronic syphilitic sores and indolent ulcer, skin infection, blood
morbidity, biliary afflictions, itching, skin ulcers,
burning sensations3 has also been evident.
Previous phytochemical studies reveales
the presence of Triterpenes: β-sitosterol, stigmasterol, Nimbidinin, Nimbidin4, Nimbin,
Azadirachtin, Flavonol
glycosides- Quercitin, Kaempferol,
Feliantriol, Salanin,
Cyclic tri and tetra sulphides. Several
investigations have proposed that this plant possess antipyretic5,
antiulcer6, immunostimulant7, antimalarial8,
hepatoprotective9, anti-inflammatory10 and antidiabetic11
effects.
Generally plants
possess many pharmacological actions since they contain numerous constituents
of active chemicals in it. Based on the above criteria this study was designed
to evaluate the activity of acetone extract on central nervous system in mice.
MATERIALS
AND METHODS:
Collection
and authentification of plant material:
The plant materials were
collected from local areas in Chennai, Tamilnadu and
the plant material was identified and authenticated by resident botanist Prof.
Dr. P. Jayaraman Plant Anatomy Research Centre
(PARC), Chennai. A voucher specimen was submitted at Annai
Veilankanni’s Pharmacy College, Chennai. Reg No; PARC/2011/860.
Preparation
of Aqueous extract12:
450gm of leaf extracted with chloroform
water by double maceration for 48hrs. The extract was filtered through mucelin
cloth. The filtrate was evaporated to dryness in vaccum
and kept in a refrigerator.
Preparation
of crude extract:
The crude extract of Azadirachta indica was freshly prepared every day by dissolving in
distilled water in order to obtain the desired concentration before the oral
administration via intra gastric tube once daily. Each animal should receive
the same volume of substance in order to avoid from the confounding error due
to different in volume.
Animals::
Wistar albino mice (25-35g) breed in central animal house
facility of the institute were used. They were housed under standard conditions
maintained on a 12 h light/dark cycle and hand free access to food and water up
to the time of experimentation. The mice were acclimatised
to the laboratory environment 1 h before the experiments. All the experiments
were conducted during the light periods (08.00-16.00 h). During the experiments
animals were free access to water only. IAEC NO: 793/03/C/CPCSEA.
Drug-chemicals::
Phenobarbitone sodium (Sigma, UK) dissolved in
saline(40mg/kg) and used accordingly. Pentazocine
(5mg/kg).
Phytochemical Screening13:
The extract was subjected to preliminary phytochemical screening by the methods previously described
by Kokate and Jayaraman J.
Acute Toxicity Study:
The procedure was followed as per OECD 423
guidelines. The extract was administered orally at a dose 2000 mg/kg body
weight to different groups of mice and observed for signs of behavioral,
Neurological toxicity and mortality 14 days.
Phenobarbitone
induced sleeping time:
Mice were given a single i.p
dose of aqueous and aqueous extract of Azadirachta indica 100 and 200 mg/kg body weight. The control
animal received the saline. Those treatments were carried out 30min before
challenging the animal with i.p injection of Phenobarbitone (40mg/kg). The latency of the loss of
righting reflex and the total sleep time [the time between the loss of recovery
of the righting reflex] were determined for each mouse as stated previously.
The mouse was considered as being awake if it could right itself (return to
upright position). Once mice right itself, it was placed on its back once more
and allowed to right a second time for confirmation14.
Hot Plate Method15:
The parameter evaluated was the latency time for paw
licking and jumping response after exposure on surface of hot plate. The
standard used was pentazocine (10mg/kg i.p). The hot plate temperature was kept at 500±
10C and the cut off time was 20 sec26.
Statistical Analysis:
The
data were expressed as mean ± standard error mean (SEM). The significance of differences among
the groups was assessed using one way analysis of variance (ANOVA). The test
was followed by Dunnett’s ‘t’-test, p values less
than 0.05 were considered as significance.
RESULTS:
Phytochemical Screening:
The
preliminary phytochemical analysis of AEAI showed
that the plant contains carbohydrates, flavanoid,
phenols, proteins, saponin but, steroids, gums and
mucilage, and tannins were absent.
Acute toxicity Study:
Acute oral toxicity studies revealed the
nontoxic nature of AEAI. There was no morbidity observed or any profound toxic reactions
found at a dose of 2000 mg/Kg p.o. which indirectly
pronouns the safety profile of the plant extract.
Phenobarbitone
Induced Sleeping Time:
Aqueous extract of Azadirachta indica leaves produce dose dependent
increase in phenobarbitone
induced sleeping time (p<0.01). The aqueous extract significantly increases the sleep duration by 0.13min,
0.15min respectively when compared to saline
treated control i.e.., 0.32 (p<0.05).
\
Group
|
Dose mg/kg (i.p) |
Mean Onset OF Sleep (min) ± SEM |
Mean Duration Of Sleep (min) ± SEM |
|
Control |
5mg/kg |
0.52 ± 0.009 |
0.32 ± 0.009 |
|
AEAI 100 |
100mg/kg |
0.59 ± 0.016 |
0.416 ± 0.026** |
|
AEAI 200 |
200mg/kg |
0.498 ± 0.007* |
0.426 ± 0.007** |
AEAI- Aqueous extract of Azadirachta indica
One way ANOVA followed by Dunnet’s test. Values are mean ± S.E.M, n=5 in each group *p<0.05,
**p<0.01, when compared to control.
Aqueous
extract of azadirachta indica leaves
produces dose dependent increase in the pain threshold time(p<0.01) when
compared to saline treated control group. The standard drug Pentazocine also
increases the pain threshold level significantly(p<0.001).
Treatment |
Dose i.p
(mg/kg) |
Mean reaction time in
seconds |
|||||
15 min |
30 min |
60 min |
90 min |
120 min |
|||
Control |
5ml/kg |
2.62 ± 0.203 |
2.95 ± 0.203 |
3.18 ± 0.244 |
3.39 ± 0.189 |
3.77 ± 0.206 |
|
Pentazocine |
5mg/kg |
4.76 ± 0.113 |
5.60 ± 0.09 |
5.30 ± 0.947 |
5.51 ± 0.102 |
5.69 ± 0.09 |
|
AEAI 100 |
100mg/kg |
3.37 ± 0.185 |
3.52 ± 0.147 |
3.76 ± 0.133 |
3.96 ± 0.123 |
4.13 ± 0.122 |
|
AEAI 200 |
200mg/kg |
3.69 ± 0.136 |
3.85 ± 0.156 |
4.03 ± 0.145 |
4.23 ± 0.149 |
4.39 ± 0.16 |
|
One way ANOVA followed by Dunnet’s
test. Values are mean ± S.E.M, n=5 in each group *p<0.05, **p<0.01,
when compared to control.
DISCUSSION:
The aqueous extract
of Azadirachta indica from
the present study produce substantial depressant effects on central nervous
system. The central action of the leaf extract was shown its ability to induce
sleep, its effects 0n phenobarbitone16 sleeping time and analgesic
activities. The extract potentiates the barbiturate sleeping time in a dose dependant
manner indicating a pharmacological action. The increase in the dose of extract
from 100 to 200mg/kg in phenobarbitone treated rats
resulted in increased duration of sleep by 0.13 min and 0.15 min. Barbiturates
are known as CNS depressants. Extract from other plants have also been observed
to potentiate the sleeping time of barbiturates and other CNS depressants. The
extract (200 mg/kg) on its own was observed to induce sleep within 10 to 15 min
of administration. This is an indication of sedative and depressant action on
the central nervous action and agrees with similar experiments in mice and rats
using other plant extracts. The depressant activity of the extract may be
attributed in part to an action on the cerebral mechanism involved in the regulation
of sleep. The result of this study also shows that the leaf extract of Azadirachta indica
induced some analgesic activity17 in rats. The plant produced
analgesia when pain was induced with heat. The extract doses (100 and 200
mg/kg) increased the time taken for paw licking and jumping response
significantly. Pentazocine (10 mg/kg) increased the
time of paw licking when compared with the extract. The analgesic superiority
of pentazocine over the extract is expected since pentazocine is a narcotic analgesic used to elevate deep
seated pain.
The data present in
our study suggest that A.indica may have some CNS depressant activity
which may be due to analgesic and Sedative-hypnotic property. This study
provides experimental support for medicinal traditional use of this plant in
nervous disorder.
ACKNOWLEDGEMENT:
The authors are
thankful to Dr. M. Senthil Kumar, Principal and Head
of the Department of Pharmaceutics Annai Veilankanni’s Pharmacy College, for his encouragement in
carrying out this work.
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Received on 07.07.2012 Accepted on 14.08.2012
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