Journal of Pharmacognosy and Phytochemistry 2020; 9(5): 485-489

E-ISSN: 2278-4136
P-ISSN: 2349-8234
www.phytojournal.com Anti-mitotic activity of aqueous leaf extracts of
JPP 2020; 9(5): 485-489
Received: 14-05-2020 Azadirachta indica A. Juss. and Simarouba glauca
Accepted: 26-06-2020
DC. On Allium cepa L. root tips
Antu Mariya Jose
Department of Botany, BCM
College, Kottayam, Kerala, India Antu Mariya Jose, Vaishnavi A and Aswathy A
Vaishnavi A
Abstract
Department of Botany, BCM
Antimitotic activity is referred as the arrest of cell multiplication in mitosis. The present study deals with
College, Kottayam, Kerala, India
the evaluation of antimitotic activity of leaf extracts of Azadirachta indica A. Juss. and Simarouba
Aswathy A glauca DC. using Allium cepa root tip cells. 50% and 25% of leaf extracts of both the plants were
Department of Botany, BCM prepared and onion root tips were exposed to each concentration separately. The root tips were then
College, Kottayam, Kerala, India squashed on microscope slides and mitotic indexes were calculated. The results indicated that the average
mitotic index of Allium cepa were considerably decreased with higher concentrations of both plant
extracts in comparison with the control. The chromosomal aberrations observed during this study are
revealing the cytotoxic effect of extracts of the selected leaves for preventing cell proliferation. The
results of present study provides information about how effective these plants in terms of their medicinal
value and also to understand why these plants can be used for developing a new anti-cancer drug.

Keywords: Antimitotic, Azadirachta indica, Simarouba glauca, Allium cepa

Introduction
The anticancer properties of plants have been recognized for centuries. The National Cancer
Institute (NCI) has screened approximately 35,000 plant species for potential anticancer
activities. Due to the advantages of medicinal plants they are of high demand. Several species
of medicinal plants have been investigated and selected for the preparation of cancer
medicines. Recently, there has been an increased scientific interest in the study of materials
from plant source as an anticancer compound [1].
The antimitotics are considered as mitosis blockers and consequently, cell death inducers.
Antimitotic agents widely used in chemotherapy, target exclusively proliferative cells and
commonly induce a prolonged mitotic arrest followed by cell death via apoptosis. As of today
Himalayan plants are a major contributor to the herbal pharmaceutical industry both of India
and other countries. Considerable works have been done on these plants to treat cancer [2].
The general principle of the mechanisms of mitosis are best and most easily studied in the
actively growing region of plants such as a shoot or root apex. Mitosis is the characteristic type
of cell division which keep the genetic continuity and integrity of organisms and preserves
their somatic chromosome value. It is also known as equational division. Since the daughter
cells formed are genetically and morphologically identical. Mitosis involves two closely
related processes, karyokinesis and cytokinesis. Karyokinesis include 5 sub stages; prophase,
prometaphase, metaphase, anaphase and telophase [3].
Azadirachta indica A. Juss commonly known as neem, or Indian lilac. It is a tree in the family
Meliaceae. It is typically grown in tropical and semitropical regions. Its fruits and seeds are the
source of neem oil. Products made from neem trees have been used in India for over two
millennia for their medicinal properties. The medicinal properties of neem have been known to
Indians since time immortal. It has been used in Ayurvedic and Unani systems of medicine
since ages. Neem products are believed to have anthelmintic, antifungal, antidiabetic,
antibacterial, antiviral, contraceptive and sedative properties [4].
A. indica has complex of various constituents including nimbin, nimbidin, nimbolide and
limonoids. Such types of ingredients play role in disease management through modulation of
various genetic pathways and other activities [5]. Accumulating evidences indicates that the
anticancer effects of neem limonoids are mediated through the inhibition of hallmark
Corresponding Author: capabilities of cancer such as cell proliferation, apoptosis evasion, inflammation invasion and
Antu Mariya Jose angiogenesis [6]. Neem leaves, natural sources of flavonoids, polyphenols, isoprenoids,
Department of Botany, BCM sulphurous and polysaccharides, play important role in scavenging the free radical and
College, Kottayam, Kerala, India subsequently arresting disease pathogenesis [7].
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Simarouba glauca DC. is commonly known as paradise tree, were taken. The numbers of dividing and non-dividing cells
dysentery bark and lakshmitaru. Its seeds produce an edible were recorded and mitotic indexes were calculated using
oil. The tree is well suited for warm, humid, tropical regions. following equation:
It is effective in treating certain diseases like diarrhea,
malaria, edema, fever and stomach upset. S. glauca has been 𝑁𝑢𝑚𝑏𝑒𝑟 𝑜𝑓 𝑑𝑖𝑣𝑖𝑑𝑖𝑛𝑔 𝑐𝑒𝑙𝑙𝑠
carried out to explore the molecular mechanisms leading to 𝑀𝑖𝑡𝑜𝑡𝑖𝑐 𝐼𝑛𝑑𝑒𝑥 (𝑀𝐼) = 𝑋 100
𝑇𝑜𝑡𝑎𝑙 𝑛𝑢𝑚𝑏𝑒𝑟 𝑜𝑓 𝑐𝑒𝑙𝑙𝑠
cancer cell death [8]. The leaves and bark have a long history
of medicinal use, right from the early centuries. It is mainly Chromosomal morphology, their changes and aberrations
used in treatment of malaria, fevers, dysentery and as an were also observed and recorded. Changes in cellular and
astringent to stop bleeding etc. The main active compounds chromosomal morphology were photographed under a light
are a group of triterpenes called quassinoids. Several of the microscope.
quassinoids found in Simarouba glauca such as ailanthinone,
glaucarubinone and holacanthone are considered the plant’s
main therapeutic constituents and are the ones documented to
be antiprotozoal, antiamoebic, antimalarial and even toxic to
cancer and leukemia cells [9].
Among the plant species, Allium cepa has been used to
evaluate DNA damages, such as chromosome aberrations and
disturbances in the mitotic cycle [10]. The ease to handle,
stable karyotype, fast response, clear mitotic phases, large
chromosome and small chromosome number are some of the
reasons that make Allium cepa suitable in cytotoxic,
antimitotic and genotoxic evaluations.
The major objective of this study is to evaluate and compare
the antimitotic activity of leaf extracts of Azadirachta indica
A.Juss. and Simarouba glauca DC. on Allium cepa root tip
cells.

Materials and Methods

Plant Collection
The leaves of both plants were collected and washed in tap
water to remove the impurities and then allowed to shade dry
under room temperature. After proper drying the leaves were Fig 1: Treatment of onion root tips with leaf extracts.
powdered separately and stored in air tight containers.
Results and Discussion
Preparation of Leaf Extracts The evaluation of anti-mitotic activity of aqueous leaf extracts
30 grams of powdered leaves of both plants were weighed and of Azadirachta indica A.Juss. and Simarouba glauca DC. in
soaked with 100ml distilled water separately and continuously onion root tips revealed that, the extracts of both leaves
stirred for 1 hour. Then it was placed in boiling water bath for showed a significant reduction in mitotic index when
30 minutes to get the aqueous extract. The extract obtained compared to that of control, and also the mitotic suppression
was then kept for 72hrs. At the end, each leaf extracts were increases with the concentration of extracts (table 1). The
filtered using filter paper, and the filtrate obtained was treatment also induced a range of chromosomal aberrations in
collected in a beaker and stored in refrigerator separately. the root tip cells of onion like nuclear lesions, prolonged
50% and 25% of aqueous extracts of both the plants were prophase, stickiness between chromosomes, laggards etc.
prepared by taking 10ml and 5ml aqueous extracts (Figure 3). When comparing the effect of both extracts, the
respectively making up to 20ml using distilled water. extract of Simarouba glauca DC.
leaves showed more antimitotic activity on Allium cepa root
Antimitotic Activity-allium cepa L. Root Assay tip cells than Azadirachta indica A. Juss. leaf extracts in both
The Allium test was performed according to the method the concentrations (figure 4). Hence the present study
described by Fiskesjo (1985) [11] and Grant (1982) [12]. Locally provides information about how effective the selected plants
available Allium cepa bulbs were washed, unscaled and in terms of their medicinal value and also to understand why
grown on sand trays for 72 hours. It was watered daily. The these plants can be used for developing a new anti-cancer
bulbs whose roots have grown to approximately 2-3cm were drug. Similar works was done in the evaluation of antimitotic
selected and washed. The clean bulbs were placed on test activity of Aplotaxis auriculata rhizomes [13] and Mukia
tubes containing aqueous extracts such that the roots were maderaspatana L. leaf extracts, [14] using Allium cepa root
immersed in the extracts. The duration of the treatment was meristematic cells and got similar results. Similar anti-mitotic
2hrs. The sprouted roots were also treated with well water, study along with phytochemical studies were conducted with
which served as control. After the treatment duration the roots polyherbal extracts by using onion root model and the results
were taken out and the root tips were hydrolyzed in 1N HCl, showed a significant reduction in mitotic index in polyherbal
stained using acetocarmine and were then squashed on extract in high concentration in comparison to standard
microscope slide covered with a cover slip. The slides were vincristine as well as in comparison to individual plant
then examined through a light microscope and photographs extracts [15].

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Table 1: Effect of aqueous extracts of Azadirachta indica A. Juss. and Simarouba glauca DC. on mitotic index of Allium cepa root tip cells.
Concentration Fields Mitotic Index (MI) Average Mitotic Index (MI)
1 87.80
Control 2 75.92 80.33
3 77.27
1 42.18
Simarouba glauca DC. (25%) 2 61.36 50.19
3 47.05
1 21.33
Simarouba glauca DC. (50%) 2 23.33 22.84
3 23.88
1 63.88
Azadirachta indica A. Juss. (25%) 2 58.88 58.97
3 54.16
1 25.75
Azadirachta indica A. Juss. (50%) 2 28.35 26.65
3 25.86

(A) (B)

(C) (D)
Fig 2: Various stages of mitotic cell divisions in Allium cepa root tip cells, (a) Prophase, (b) Metaphase, (c) Anaphase, (4) Telophase

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(A) (B)

(C) (D)
Fig 3: Chromosomal aberrations observed in Allium cepa root meristem cells treated with aqueous extracts of Azadirachta indica and
Simarouba glauca. (a) Nuclear lesions, (b) Prolonged prophase, (c) Stickiness between chromosomes, (d) Laggard.

inhibitory and mitodepressive effects on the cell division of

Allium cepa L. meristems. Hence these plants can be used for
their anti-cancer properties. Further studies have to be
conducted to obtain more detailed mechanism of action of
plant material in view of its antimitotic activity.

Acknowledgement
The authors are greatful to Dr. Peter K. Mani, Head Of the
Department of Botany, B C M College Kottayam for his
constant support throughout this work as part of completion
of B. Sc degree.
Fig 4: Antimitotic activity of leaf extract of Azadirachta indica
A.Juss. and Simarouba glauca DC. on mitotic index of onion root Reference
tips. 1. Avni G Desai, Ghulam N Qazi, Ramesh K Ganju,
Mahmoud El-Tamer, Jaswant Singh, Ajit K Saxena et al.
Conclusion Medicinal Plants and Cancer Chemoprevention. Current
The present study was an attempt to evaluate the antimitotic Drug Metabolism. Bentham Science Publishers LTD.
activity of leaf extracts of Azadirachta indica A. Juss. and 2008; 9(7):581-591.
Simarouba glauca DC. on Allium cepa L. root tip cells. The 2. Umadevi M, Sampathkumar KP, Debjit Bhowmik,
results revealed that, the extracts of both leaves showed a Duraivel S. Traditionally Used Anticancer Herbs in India.
significant reduction in mitotic index of root tips when Journal of Medicinal Plant Studies. 2013; 1(3):56-74.
compared to control, and also the mitotic suppression 3. Mitsuhiro Yanagida. The Role of Model Organisms in
increases with the concentration of extracts. On comparison the History of Mitosis Research. Cold Spring Harb
of antimitotic activity of two selected plants, the leaf extracts Perspect Biology. 2014; 6(9):1-14.
of Simarouba glauca DC showed marked antimitotic property 4. https://scholar.google.com/scholar?hl=en&as_sdt=0%2C
than that of Azadirachta indica A. Juss. Certain chromosomal 5&q=D.+P.+Agrawal%2C+medicinal+properties+of+nee
aberrations were also observed in the higher concentrations m&oq=#d=gs_qabs&u=%23p%3D_2VoTdnphyUJ
(50%) of both plant leaf extracts. Thus the present study 5. Mohammed A Alzohairy. Therapeutic Role of
suggested that the aqueous extracts of leaves of Azadirachta Azadirachta indica (Neem) and Their Active
indica A. Juss. and Simarouba glauca DC. has significant Constituents in Disease Prevention and Treatment.
~ 488 ~
Journal of Pharmacognosy and Phytochemistry http://www.phytojournal.com

Evidence-Based Complementary and Alternative

Medicine. 2016; (11):1-11.
6. Nagini S, Priyadarsini RV. Azadirachta indica (Neem)
and Neem Limonoids as Anticancer Agents-Molecular
Mechanisms and Targets. Perspective in Cancer
Prevention-Translational Cancer Research. Springer,
Newdelhi, 2014, 45-60.
7. Dharmendra Kumar Yadav, Yogesh P Bharitkar, Kasturi
Chatterjee, Monisankar Ghosh, Nirup Bikash Mondal,
Snehastika Swarnakar et al. Importance of Neem leaf-An
insight into its role in combating diseases. Indian Journal
of Experimental Biology. 2016. 54(11):708-718.
8. Asha Jose, Elango Kannan, Palur Ramakrishnan, Anand
Vijayakumar, SubbaRao Venkata Madhunapantula.
Therapeutic Potential of Phytochemicals Isolated from
Simarouba glauca for Inhibiting Cancer: A Review.
Systematic Reviews in Pharmacy. 2019; 10(1):73-80.
9. http://tropical.theferns.info/viewtropical.php?id=Simarou
ba+glauca
10. Daniele Morais Leme. Allium cepa in environmental
monitoring: A review on its application. Mutation
Research Fundamental and Molecular Mechanisms of
Mutagenesis. 2009; 682(1):71-81.
11. Geirid Fiskejo. The Allium test as a standard in
environmental monitoring. Heriditas. 1985. 102: 99-112.
12. Grant, W. F. Chromosome aberration assays in Allium.
Mutation Res. 1982; 99:273-291.
13. Shalini R, Velavan S. Evaluation of Antimitotis Activity
of Aplotaxis auriculata Using Allium cepa root
Meristamatic Cells. Indian Journal of Applied Research.
2017; 7(12):315-317.
14. Kavitha M, Srinivasan PT, Renuga, Jayakumar LV.
Evaluation of Antimitotic Activity of Mukia
maderaspatana L. Leaf extracts in Allium cepa Root
Model. International Journal of Pharmacy Research and
Technology. 2014; 4(1):65-68.
15. Anntoniya Rintelen, Kelvin Bucktowar, Ranajit DT.
Evaluation of Phytochemical and Antimitotic Activity of
Poly Herbal Extract by Using Onion Root Model.
International Research Journal of Pharmacy. 2017;
8(4):44-49.

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