melia azedarach as a immunomodulatory plant

2. Introduction to medicinal plants.
Medicinal plant Melia azedarach L.
Immunomodulation and immunomodulators
In- vitro & in-vivo tests for evaluation of
immunomodulators
MTT assay
Thesis work plan

3.  India has a wealth of medicinal plants most of which have
been traditionally used in Ayurveda, Unani & Sidhaa.
 There are about 6000-7000 medicinal plant species(NMPB)in
India, with concentrated spots in the region of Eastern
Himalayas, Western Ghats and Andaman & Nicobar Island.
 India is the largest producer of medicinal herbs and is
called the botanical garden of the world.
 25% of modern pharmaceutical drugs contain plant
ingredients.

4. herbals are relatively inexpensive
need for alternative treatments for drug-
resistant pathogens
expanding contact and growing respect for
foreign cultures, including alternative systems
of medicine
High commercial value of medicinal plants.

5. phenolics 45%
alkaloids 18%
terpenoids & steroids
27%
others 10%

6. Ref:International journal of
herbal medicine

7. Common name:chinaberry, White cedar,
bakain, syringa, ghora neem, umbrella tree etc.
 native to Southeast Asia and northern
Australia
 deciduous tree growing to 50 ft. in height and
2 ft. in diameter.
 Plant Parts Used : Leaf, flower, root, young
branches, fruit and bark.

8. Kingdom: Plantae
Phylum: Magnoliophyta
Class: Magnoliopsida
Order: Sapindales
Family: Meliaceae
Genus: Melia
Species: azedarac

9. Medicinal activity Targeted organism or
disease
Extracts Basic Reference
Larvicidal &
anti- oviposition
Pediculus humanus capitis Leaf & Fruit Carolina et al, 2008, ,
Carolina et al. 2004,
Cori a et al 2008,
Pediculicidal &
Ovicidal
Pediculus humanus capitis Fruit Caprinella et al,
2003-2007
Antifeedant Pseudaletia unipunctata
Trachiaplusia ni
Spodoptera eridania
Seed
Seed
Fruit
Akhtar et al 2008
Carpinella et al.,
2002

10. Medicinal activity Targeted organism or
disease
Extracts Basic Reference
Rodenticidal Albino rat Seed
Leaves
Roop et al 2005,
Keshri et al 2003,
Keshri et al, 2004
Fungicidal Aspergillus flavus
Diaporthe phseolorum
Schlertina sclertiorum
Seed Carpinella et al,
1999-2003-2005
Antibacterial Proteus mirabilis
Shigella flexeneri
Staphyloccocus aureus
Bacillus subtilis
Leaf Abdul Viqar et al.,
2008
Anti helminthal Haemonchus contortus Seed, Leaf Maciel, et al, 2006

11. Medicinal activity Targeted organism or disease Extracts Basic Reference
Antiviral Foot and mouth Disease
Herpes simplex,(VSV) ,
(HSV)(VSV), (HSV1)
Ocular Herpes simplex
Leaf
Leaf
Leaf
Leaf
Wachsman et al.,
2008
Wachsman et al.,
1982
Alche, L. et al. 2002
Pifarri, et al. 2002
Antiprotozoal Trichomonas vaginalis Seed,
Leaf
Lee et al., 2007
Anti hatchability Erias vitella Seed Gajmer et al, 2001

12.  Immunomodulation is the the alteration of immune response
which may increase or decrease the immune responsiveness.
 The immune response is how our body recognizes and
defends itself against bacteria, viruses, and substances that
appear foreign and harmful

14.  An immunomodulators may be defined as a substance,
biological of synthetic, which can stimulate, suppress or
modulate any of the components of the immune system
including both innate and adaptive arms of the immune
response
 2 types of immunomodulator
1. immunostimulators, are substances (drugs and nutrients)
that stimulate the immune system by inducing activation or
increasing activity of any of its components.
2. An immunosuppressant is any agent that weakens the
immune system.

15. 1) In vitro
a) lymphocyte proliferation assay
Quantitates the response of T cells to a variety of stimuli
including mitogens, alloantigens and specific recall antigens.
b) Jerne’s plaque assay
an assay that enumerates individual antibody-forming
cells

16. c) IL-1 and TNF – alfa production by
macrophages in response to
lipopolysaccharides
d) quantitation of the expression of cell
activation markers on cell surface viz.
CD25,CD69,CD86 by fluorescent activated
cell sorting(FACS).
e) augmentation or inhibition of NK cell activity
in absense or presense of IFN- gama.

17. IN- VIVO:
a) estimation of specific antibody profile and
titer to specific response.
b) Delayed Type Hypersensitivity (DTH)
response to specific antigen
c) spleen foci (colony formation) assay for
immunostimulant activity

18. d) Induction/progression of autoimmune disease in
experimental animal models for immunosuppresant
activity.
e) Prevention of allograft or xenograft rejection.
f) Evaluation in rodent malaria and japanese
encephalitis virus models for immunostimulant
activity.

19.  MTT assay is a laboratory test and a standard colorimetric assay (an assay
which measures changes in color) for measuring cellular proliferation (cell
growth).
 It can also be used to determine cytotoxicity of potential medicinal agents
and other toxic materials.
 Yellow MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium
bromide, a tetrazole) is reduced to purple formazan in the mitochondria of
living cells.

 A solubilization solution (usually either dimethyl sulfoxide, an acidified
ethanol solution, or a solution of the detergent sodium dodecyl sulfate in
dilute hydrochloric acid) is added to dissolve the insoluble purple formazan
product into a colored solution.

20. Fig: A microtiter plate after an
MTT assay increasing amount of
cells resulted in increased purple
color

21. Establishment of chicken lymphocyte cell line
culture
In – vitro test for evaluation of immunomodulator
Expression analysis of cell cytokines

22. Making of aqueous/methanol extract from M.
azedarach leaf.
chicken lymphocyte cell line culture
determination of max. non cytotoxic dose of
leaf extract by MTT assay

23. performing lymphocyte proliferation assay at max.
non cytotoxic dose
expression analysis of cell cytokines (IL-2,IL-6)

24. • Scientific validation of this plant as an
immunomodulator.