Phytochemical and antibacterial activity was tested by using leaf extract

1. WELCOME
PRESENTED BY,
R. SIVA DHARSHINI,
Avinashilingam Institute for home
science and higher education for
women, Coimbatore- 43, Tamilnadu,
India.

2. PHYTOCHEMICAL
ANALYSIS AND
ANTIBACTERIAL
ACTIVITY OF
LEAVES EXTRACT
FROM

3. INTRODUCTION
Sustainable agriculture aims at reducing the
incidence of pests and diseases to such a degree that they do not
seriously damage crops without upsetting nature’s balance.
One of the aims of sustainable agriculture is to
rediscover and develop strategies whose cost and ecological side-
effects are minimal.
The use of synthetic pesticides has undoubtedly
resulted in achievement of green revolution in different countries
through increased crop production.
However, in recent years there has been
considerable pressure on consumers and farmers to reduce or eliminate
synthetic pesticides in agriculture. This concern has encouraged
researchers to look for better alternatives to synthetic pesticides.

4. Even though pharmacological industries have
produced a number of new antibiotics in the last three decades,
resistance to these drugs by microorganisms has increased.
In general, bacteria have the genetic ability to
transmit and acquire resistance to drugs, which are utilized as
therapeutic agents .
Such a fact is cause for concern, because of the
number of patients in hospitals who have suppressed
immunity, and due to new bacterial strains, which are multi-
resistant.
Consequently, new infections can occur in hospitals
resulting in high mortality.

5. For a long period of time, plants have
been a valuable source of natural products for
maintaining human health, especially in the last
decade, with more intensive studies for natural
therapies.
According to World Health
Organization, medicinal plants would be the best
source to obtain a variety of drugs.
About 80% of individuals from
developed countries use traditional medicine, which
has compounds derived from medicinal plants.
Therefore, such plants should be
investigated to better understand their properties,
safety and efficiency .

6. To concern both,
alternatives to synthetic pesticides to invade
pest population and
development of compounds from medicinal
plants to derive as a medicine and antibiotics
without harm effects.
So, We analysed the phytochemicals and
antibacterial activity of one of the medicinal
plant’s leaves extract, Pongamia glabra.

7. Pongamia glabra
Kingdom: Plantae
Order: Fabales
Family: Fabaceae
Genus: Pongamia
Species: glabra

8. Pongamia glabra is a species of tree in
the pea family, Fabaceae, native in tropical and
temperate Asia including parts of Indian
subcontinent, China, Japan, Malesia,
Australia and Pac
ific islands.
It is often known by the
synonym Pongamia pinnata .
Common names include Indian
beech, Pongam oiltree, karanj (Hindi),
honge (
in Kannada), pungai (
in Tamil), kānuga (in Telugu), karach (
in Bengali), naktamāla (in Sanskrit).

9. In this age old practice, fresh pungam leaves
were placed as layers in between the gunny bags
arranged one above other in storerooms.
These leaves are acted as repellent against
angumois grain moth (Sitotroga cerealella) and rice
weevils (Sitophilus oryzae).
The strong odour released from pungam
leaves avoided pest attack.
Some farmers placed these pungam leaves
directly in the gunny bags and stored the grains.

10. The seed oil being antiseptic,
antipruritic and analgesic in properties, is used
with great benefit.
The seed powder is effective as
nasal therapy to relieve the phlegm in chronic
sinusitis and kapha diseases.
It is beneficial in anorexia, piles,
worm infestations, and flatulence and liver
diseases.
Karanja is a good blood purifier.

11. Materials and Methods:
PROCESSING OF PLANT SAMPLES:
The leaves of the plants are properly washed in tap water and then
rinsed in distilled water. The rinsed leaves are dried
in shade until it dried. The dried leaves of each plant are pulverized,
using a sterile electric blender, to obtain a powered form. The
powdered form of these plants is stored in airtight plastic
bag.
PREPARATION OF EXTRACT:
Ten grams of dried plant material was extracted with 100 ml of
acetone and choloroform solvents kept on a rotary shaker for 24 hour.
Thereafter, it was filtered. The filterate was collected and the solvent
was evaporated to make the final volume one-fifth of the original
volume . It was stored in airtight bottles for further studies.

12. ANTI BACTERIAL ACTIVITY:
BACTERIA:
The bacterial strains are identified strains and were obtained. The
bacterial strains studied are Bacillus subtilis, Streptococcus aurelus,
Klebsiella pneumoniae and Escheria coli.
ANTIBACTERIAL ASSAY:
The antibacterial activity of plant species was evaluated by agar
well diffusion for solvent extract using Mueller Hinton Agar No. 2
and Agar agar medium for the assay. Petriplates containing 20 ml
Muller Hinton medium were seeded with 24 hours culture of
bacterial strains separately. Wells were cut and 20 µl of the sample
(acetone and chloroform extracts) were added. The plates were then
incubated at 37° C for 24 hours. The antibacterial activity was
assayed by measuring the diameter of the inhibition zone formed
around the well (NCCLS, 1993). Chloramphenicol was used as a
positive control. DMSO4 was used as a negative control.

13. QUALITATIVE PHYTOCHEMICAL
SCREENING:
The following tests were performed on different extracts to detect various
phytoconstituents using standard protocols (Preethi and Piramu, 2013).
Detection of alkaloids
Wagner’s test: One mL of filtrate was taken, few drops of Wagner’s reagent was
added to the side of the test tube. A reddish brown precipitate confirms the
positive results.
Detection of carbohydrates
Fehling’s test: One mL of the filtrate was boiled in water bath with 1mL each of
Fehling’s solution A and B. Brick red precipitate was formed, that indicates the
presence of sugar.
Detection of saponins
Foam’s test: The extract (50mg) was diluted with distilled water and make up to
20mL. The suspension was shaken in a graduated cylinder for 15mins. A two cm
layer of foam indicates the presence of saponins.

14. Detection of phenolic compounds and flavonoids
Lead acetate test: The extract of 50mg was dissolved in distilled water
and, 3mL of 10% lead acetate solution was added. A bulky white
precipitate indicates the presence of flavonoid compounds.
The extract of 1ml was dissolved in distilled water and few drops of
ferric chloride added to it. A dark blue precipitate indicates the
presence of polyphenols.
Test for tannins
To one ml test solution, 2-3 ml of basic lead acetate solution was
added. White precipitate was formed. It indicates the presence of
tannins.
Test for Steroids:
To 4ml of extract, 0.5ml of acetic anhydride and 0.5ml of chloroform
and then add few drops of sulphuric acid slowly. A green bluish colour
indicates the presence of steroids.
Test for Reducing Sugar:
The mixture of few drops of Benedict’s reagent with one ml of extract
was soaked in the sterile stick and it in bunsen flame. Apprearance of

15. OUTLINE OF THE STUDY
 We did the anti bacterial activity by using Agar well
diffusion method using Muller Hinton Agar medium.
Wells
 A- Acetone Extract
 B- Chloroform Extract
 C- Chloronphenicol(Positive Control)
 D- Dimethyl Sulfoxide(Negative Control)
 The tests were performed on different extracts
to detect various phytoconstituents using standard
protocols (Preethi and Piramu, 2013)
STUDY OUTLINE

16. RESULT AND DISCUSSION
ANTIBACTERIAL ACTIVITY:
Antibacterial activity assay of all the two solvent extracts
revealed that acetone and chloroform extracts showed
significant activity against both
phytopathogenic and human pathogenic bacteria. The plant
extracts were more active against the Gram- negative
bacteria than against the Gram- positive bacteria. Acetone
extract showed highly significant activity when compared
with chloroform against plant pathogenic bacteria (table 1).
RESULT AND DISCUSSION

17. GRAM POSITIVE BACTERIA
Staphylococcus aurelus Bacillus substilis

18. GRAM NEGATIVE BACTERIA
Klebsiella pnemoniae Escheria coli

19. Bacterial Strains Zone of inhibition(mm)
Acetone Chloroform Chloron
phenicol
DMSO4
Gram Positive Bacteria
Bacillus substilis 0 0 10 0
Staphylococcus aureus 0 0 12 0
Gram Negative Bacteria
Escheria coli 9 0 1
1
0
Klebsiella pnemoniae 12 8 13 0

20. 0
2
4
6
8
10
12
14
Bacillus Substilius Staphylococcus aurelus Escheria coli Klebsiella pnemoniae
Acetone
Chloroform
Chloronphenicol
DMSO4

21. The polarity of the solvent seems to play an important role in
exhibiting potential antibacterial activity.
Here, acetone and chloroform extracts of Pongamia glabra
showed remarkable activity against some medically
important bacterial strains.
In addition such results justify the traditional use of
Pongamia glabra. Further it also supports some of the
phytochemical and pharmacological investigation of this
plant carried by many researchers. This is in agreement with
previous reports that plant extracts are more active against
Gram-positive bacteria than against Gram-negative bacteria.
Bacillus subtilis and Staphylococcus aureus was the most
susceptible bacteria amongst all the bacterial strains
investigated in the present work.
The results suggest that traditional folk medicine could be
used as a guide in our continuing search for new natural
products with potential medicinal properties.

22. QUALITATIVE PHYTOCHEMICAL
SCREENINGSCREENING
Table 2 Shows that the phytochemical constituents of
Acetone, and chloroform extracts of Pongamia glabra. The
phytochemical screening of the extracts revealed the
presence of Flavonoids, alkaloids, saponins, tannins, and
polyphenols.
Saponins were present in chloroform extract whereas, the
acetone extract showed negative result for saponins.
In the case of flavonoids, acetone extract showed
positive results.
Both the extracts showed the absence of proteins.
The reducing sugar were also absent in chloroform
extract and the acetone extract shows negative.
QUALITATIVE
PHYTOCHEMICALS

23. Phytochemicals Plant Extracts
Acetone Chloroform
- +
+ -
- +
+ -
- -
- -
+ +
- -
- -
- +
Alkaloids
Flavonoids
Saponins
Tannin
sAmino acids
Proteins
Steroids
Reducing Sugars
Carbohydrates
Polyphenols

24. PHYTOCHEMICALS ANALYSIS
FOR
ACETONE
EXTRACT
FOR
CHLOROFORM
EXTRACT

25. The glycosides present in acetone
extract and amino acids were absent
in all extract
. The acetone extract shows the
positive result of tannins.
In the case of alkaloids the
chloroform extract shows positive
whereas the acetone extracts shows
negative.
All extracts showed negative results
for amino acid test. The results
obtained in the present study also

26. 0.3
0.2
0.1
0
0.4
1
0.9
0.8
0.7
0.6
0.5
Acetone
Chloroform

27. Phytochemical analysis conducted
on the plant extracts revealed the
presence of constituents which are
known to exhibit medicinal as well
as physiological activities .
Analysis of the plant extracts
revealed the presence of
phytochemicals such as phenols,
tannins, flavonoids, saponins,
glycosides and alkaloids.