International Journal of Research ISSN NO:2236-6124

PHYTOCHEMICAL SCREENING, IN VITRO ANTIOXIDANT, ANTIMICROBIAL

AND ANTI-INFLAMMATORY ACTIVITIES OF AQUEOUS EXTRACT OF MELIA
AZEDARACH

G. VIDHYASRI AND G. PRASANNA*

PG and Research Department of Biochemistry, Sengamala Thayaar Educational Trust

Women’s College (Autonomous), Sundarakkottai, Mannargudi - 614 016, Tamil Nadu,
India. (Affiliated to Bharathidasan University, Tiruchirappalli, Tamil Nadu, India).
*Corresponding Author

ABSTRACT
The aim of the present study is to examine the phytochemical, In vitro antioxidant
antimicrobial and anti-inflammatory activities of the aqueous leaf extract of Melia azedarach.
Phytochemical and proximate analysis was done using standard methods. The leaf extract of
the plant were tested against [Link], [Link], [Link] and [Link] agar diffusion method.
The free radical scavenging activities of the leaves were evaluated by various in vitro
methods. The anti inflammatory activity of plant was done by protein denaturation method.
Phytochemical, proximate analysis results showed that the presence of various
phytochemical. The proximate composition shows that the plant has high carbohydrate
(25.4%), moderate concentration of protein (16.4%) and low concentration of fat (3.2 %).
Alkaloids were found to be the most abundant one followed by phenols and flavonoids while
tannins and saponins were least in concentration. All tested concentrations of M.
azedarach extract showed significant antibacterial and antifungal activities and a high anti-
inflammatory activity compared with diclofenac as drug. Our investigation pointed that M.
azedarach could be considered as a good medicinal agent. In our knowledge aqueous extract
of Melia azedarach showed a potential antioxidant, antimicrobial and anti inflammatory
activity because of its high secondary metabolites.

INTRODUCTION
Medicinal plants plays an important role in supporting the health care and are used
since ancient times for the remedies for human diseases as they contain numerous
phytochemicals component with high medicinal value and are considered to be more natural
and safe when compared to synthetic drugs (Kannabiran et al ., 2009).Who estimated that
80% of rural population depends on locally available herbs for their primary health care. In
traditional and allopathic medicine the use of plant and its products has a long history (Dubey
et al., 2007). Pharmacological properties of the plant species are due to the presence of
various secondary metabolites like glycosides, saponins, flavonoids, steroids, tannins,
alkaloids, tirpenes which is therefore, should be utilized to combat the disease causing
pathogen (Kamali&Amir, 2010). With the advancement in Science and Technology,
remarkable progress has been made in the field of medicine with the discoveries of many
natural and allopathic drugs (Preethi et al., 2010).

Melia azedarach Family (Meliaceae) is the best known species and also known as
cinnamon or chinaberry, is a medium-sized species. It is widely cultivated in Asia and South
of Latin America being quite adapted to the cold weather Their tree can reach more than 40
meters high and is hardy and draught resistant (Seth., 2004). Leaves are alternate, 20-40 cm
long, bipinnate or occasionally tripinnate. Leaflets 3-11, serrate, dark green on the upper
surface and paler underneath. They produce a pungent odour when crushed. Inflorescence a
long, axillary panicle up to 20 cm long. Flowers are purple and fragrant, numerous on slender

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International Journal of Research ISSN NO:2236-6124

stalks, white to lilac. Its various parts have antihelmintic, antimalarial, cathartic, emetic and
emmenagogic properties and are also used to treat skin diseases (Pokhrel et al., 2015). It has
also been used as an abortifacient, an antiseptic, antioxidant a purgative, a diuretic, and an
insect repellent. It is used for generally healing arthritis, rheumatism, for Pulmonary, stomach
troubles, diarrhoea and dysentery. It is also used as vermifuges, to treat, cutaneous,
subcutaneous parasitic infection and for menstrual cycle problems.

MATERIALS AND METHODS

Collection of plant and Preparation
Melia azedarach leaves were collected from STET garden, Mannargudi, Thiruvarur
district. The leaves were washed with the distilled water for about two to three times, and
then, it was cut into small pieces and shade dried for several days. The leaves were grounded
using motor and pestle and stored in an airtight container. The powered leaves were mixed
with water to prepare aqueous extract.

Qualitative phytochemical analysis

The qualitative test was performed in the extract for the identification of various
phytochemical constituents as per standard procedures (Sofowora, 1993).

Proximate Analysis
Proximate analysis such as moisture content, Ash content (AOAC, 2000), crude fibre,
crude lipid (AOAC, 1990), Protein (Lowry’s method 1951) and Carbohydrate (Dubois et al.,
1956) was determined.

Quantitative analysis
The extract was subjected to quantitative analysis for the determination of secondary
phytochemical constituents Determination of total alkaloids (Obadoni and Ochuko, 2001),
Total phenolic content (Kaur and Kapoor (2002).) Total flavonoids content (Chang et al.,
(2002). Estimation of total tannins (Siddhuraju and Manian (2007). Estimation of total
saponins (Makkar et al., (2007).

In vitro Antioxidant Activity

Antioxidant activity of the extract was determined by various methods such as DPPH
radical scavenging activity (Mensor et al., 2001), Superoxide radical scavenging activity
(Awah et al., 2010), Hydrogen Peroxide Scavenging Activity (Ruch et al., 1989) and Total
antioxidant capacity (Habib et al., 2014).

Screening of Antimicrobial Activity

In-vitro antimicrobial screenings were carried out under laboratory conditions, for this
various micro organism were collected from microbiology laboratory, STET, Mannargudi
with bacterial strain of [Link] and [Link] and fungal strain of Aspergillus niger and
candidas albicans . All the microorganisms were cultured on recommended cultural medium
and transfer & maintained on agar broth for O/N. Antimicrobial activity of plant Melia
azedarach have been carried out by using disc diffusion method (Kavanagh, 1972). The
inhibitory effect of extract was compared with the standard antibiotics penicillin and
mycostatin against bacteria and fungi respectively.

Anti inflammatory activity

Anti inflammatory activity of the plant extract was determined by invitro method
such as protein denaturation (Mizushima et al., 1968, Elias & Rao, 1988).

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RESULT AND DISCUSSION

Table 1. Preliminary Phytochemical Analysis of extracts of the leaf of Melia azedarach

S. No Constituents Aqueous
1 Alkaloids +
2. Flavonoids +
3 Phenols +
4 Steroids +
5 Tannins +
6 Saponins +
7 Terpenoids +
8 Cardiac glycosides +
9 Sugar +
10 Aminoacids +

The phytochemical analysis of Melia azedarach was shown in table 1. Preliminary

investigations revealed the presence of alkaloids, flavonoids, phenols, steroids, tannins,
saponins sugar, aminoacids, terpenoids and cardiac glycosides. Phytochemicals in the plants
are non-nutritive chemicals that have protective or disease preventive properties. The
presence of phytochemicals in Melia azedarach possesses varying degrees of disease
preventive antioxidants and antimicrobial molecules. Hence, it is termed as health protective
agent. In addition, these phytochemicals acts as best antioxidants and protect the cells against
free radical damage (Suresh and Nagarajan, 2011).

Proximate Composition
The results of proximate composition of Melia azedarach is presented in Table [Link]
proximate analysis showed the moisture content of Melia azedarach to be14.8% and the
result indicated that shelf life of the fresh plant was low which lead to spoilage due to its
susceptibility to microbial attack. Moisture content is among the most vital and mostly used
measurement in the processing, preservation and storage of food (Onwuka, 2005).

Table 2: Proximate composition of Melia azedarach leaves

[Link]. Parameters Value (%)

1. Moisture content 14.8

2. Total Ash
9.8
3. Crude Fibre
8.3
4. Crude lipid
3.2
5. Protein
16.4
6. Carbohydrates
25.4

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Ash content of 9.8 % dry matter was obtained as a result for Melia azedarach. Ash in
food contributes the residue remaining after all the moisture has been removed as well as the
organic material (fat, protein, carbohydrates, vitamins, organic acid etc) have been
incinerated at a temperature of about 500oC. Ash content is generally taken to be a measure
of the mineral content of the original food (Onwuka, 2005). Crude fibre in food or plant is an
indication of the level of non-digestible carbohydrate and lignin. The crude fibre obtained for
Melia azedarach was 8.3% .This low level is considered appropriate, because it aids
absorption of glucose and fat. Although crude fibre enhances digestibility, its presence in
high level can cause intestinal irritation, lower digestibility and decreased nutrient usage
(Oladiji et al., 2005).

Crude fibre is made up largely of cellulose together with a little lignin which is
indigestible in human. The crude lipid content obtained for Melia azedarach was 6.2%. Lipid
provides very good sources of energy and aids in transport of fat soluble vitamins, insulates
and protects internal tissues and contributes to important cell processes (Pamela et al., 2005).
More so, it is good to add lipid (fat) to most of our diets, because many body functions
depend on lipids. The crude protein of Melia azedarach was 16.4 %. The plant is a moderate
source of protein. According to Pamela et al., (2005), proteins from plant sources have lower
quality but their combination with many other sources of protein such as animal protein may
result in adequate nutritional value. The carbohydrate content of M. azedarach was 25.4 %.

Table 3: Quantitative Phytochemical composition of Melia azedarach leaves

S. No. Parameters Value (mg/100g)

1. Alkaloids 8.45 ± 0.17

2. Flavonoids 3.8 ± 0.03

3. Phenols 7.46 ± 0.01

4. Tannin 3.2 ±0.05

5. Saponin 1.50 ±0.01

The data of quantitative determination of secondary metabolites as tabulated in table 3

shows that among the five groups of phytochemicals determined from the leaves of Melia
azedarach, alkaloids were found to be the most abundant one followed by phenols and
flavonoids while tannins and saponins were least in concentration. Phenols and tannins have
ability to scavenge ROS and have antioxidant properties .Various reports indicate that regular
flavonoids may trim down the risk of several chronic diseases including neurodegenerative
diseases, atherosclerosis, and cancer (Mahesh et al., 2013). The flavonoids also reported to
have antiviral, anti allergic, anti platelet and anti inflammatory properties.

Antioxidant Activity
The result of the present study showed that the aqueous leaf extracts of Melia
azedarach exhibited the greatest anti-oxidant activity .The high scavenging property of the
medicinal plants may be due to hydroxyl groups existing in the phenolic compounds. It is
reported that phenols are responsible for the variation in the antioxidant activity of the plant

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(Cai et al., 2004). They exhibit antioxidant activity by inactivating lipid free radicals or
preventing decomposition of hydroperoxides into free radicals (Pitchaon et al., 2007). It is
also reported that antioxidant activity of phenolic compounds is mainly due to their redox
properties (Rahman, 2007).Vegetables contain a wide variety of biologically active, non-
nutritive compounds known as phytochemicals. This is often attributed to the antioxidants
such as vitamin C, E, carotenoids, lycopenes and flavonoids that prevent free radical damages
(Oomah & Mazza, 2009). These phytochemicals impart health benefits beyond basic
nutrition.
Table 4: Antioxidant activity of Melia azedarach leaves

S. No Concentration DPPH SOD % H2O2% TAC Ascorbic

(%) (%) acid (%)
1. 100 26±1.3 15.4±0.77 18.9±0.7 25.7±0.2 24.8±0.25

2. 200 35.7±1.7 24.7±1.2 22.3±0.6 31.1±0.5 30.7±0.6

3. 300 43.4±0.9 29.1±0.6 26.4±0.9 35.8±0.3 35.15±0.7

4. 400 50.5±2.5 32.5±1.4 31.2±0.6 38.8±0.4 40.6±0.8

Antimicrobial analysis
In the present study, the aqueous extract of leaves of M. azedarach showed
antimicrobial activity against B. subtilis and E. coli from bacteria, A. niger and C. albicans
from fungi. The finding of this study revealed that the leaves of M. azedarach have broad
spectrum antibacterial potential. The results obtained correlated well with the report of Sen
and Batra, 2012 in which the ethanol extract of leaves of M. azedarach showed antimicrobial
activity against clinical pathogens such as B. cereus, S. aureus, E. coli, P. aeruginosa, A.
flavus and A. niger. Majeed (2013) examined the antimicrobial activity of M. azedarach
leaves against S. aureus, Klebsiella sp., E. coli and P. aeruginosa. He reported that the extract
showed better activity against S. aureus at a concentration of 100μg/ml compared to other
microbes used in their study. The antimicrobial analysis (Table 5.) showed a remarkable
activity against the bacterial and fungal pathogens with aqueous extract of Melia azedarach.
The maximum activity compared to the control shows the potential of the plant and is an
indicator for determining the significance of the activity against the pathogens. The overall
antibacterial analysis reveals maximum against the B. subtilis and minimum activity was
noted against the [Link].

Table 5: Antimicrobial activity of Melia azedarach leaves

S. No Microbes control Standard (Penicillin Extract
and mycostatin)
1. Escherichia coli - 18.0±0.69 8.8±0.3

2. Bacillus subtilis - 22.8±0.6 13.7±0.8

3. Aspergillus niger - 12.3±0.18 10.5±0.3

4. Candidas - 11.1±0.7 9.3±0.08

albicans

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Anti-inflammatory activity
The protein denaturation method was used for the in vitro anti-inflammatory activity
of the aqueous extract of the Melia azedarach which was presented in Table 6. % inhibition
of protein denaturation activity was exhibited on the basis of concentration dependent
manner. The % inhibition of protein denaturation was 22.8% for 100µg/ml and 38.2% for
500µg/ml. Sodium diclofenac was used as a standard. During inflammation the lysosomal
enzymes is released which produced a variety of disorders and these enzymes is said to be
related to acute or chronic inflammation. The diclofenac drugs act either by inhibiting these
lysosomal enzymes or by stabilizing the lysosomal membrane. The aqueous extracts of leaves
Melia azedarach showed biphasi effects on protein denaturation method.

Table 6: Anti inflammatory activity of Melia azedarach leaves

S. No. Concentration % inhibition of protein

(μg/ml) denaturation
1. 100
22.8±0.3
2. 200
25.1±0.4
3. 300 28.2±0.7

4. 400 34.3±1.2

5. 500 38.2±0.4

6. Standard 74.6±0.14
(Diclofenac)

On the basis of the results obtained in the present study, it is concluded that aqueous
extract of Melia azedarach leaves has potent anti-inflammatory and anti microbial activities.
Thus the Melia azedarach extract may be attributed to the presence of phenolic compounds
and flavonoids etc., Therefore, further investigation is needed to isolate and identify the
active compounds present in the plant extract and its efficacy.

Acknowledgement

The authors are grateful to the Management of Sengamala Thayaar Educational Trust
Women s College (Autonomous), Sundarakkottai, Mannargudi for their encouragement and
support.

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