Research and Basic information about Medicinal Plants.

1. PHYTOCHEMICAL INVESTIGATION AND THE
STUDY OF DRUG POTENTIAL OF PHYTO
COMPOUND IN Nicotiana tobaccum
Presented by
Mr. Halavath. RAMESH
Dept. of Chemistry
Loyola College, Chennai.
1

2. • Introduction
• Experimental section
• Results and discussion
• Summary and conclusions
• References
• Acknowledgements
2
CONTENT

3. 1. Phytochemicals are chemical compounds produced by plants.
2.Challenges in the field phytochemistry including isolating specific
compounds and determining their structure and identifying what specific
Phytochemical is primarily response in biological activity.
3. In India Herbal medicines play an important role in the treatment of
various diseases by trational methods practiced such as Ayurveda , Unani
and Siddha.
4. Over 2,48,000 species of the higher plants have been identified 12,000
plants are known to have medicinal properties.
5. The knowledge of pharmacology in essential for understanding action of
drugs on animals and human system.
6. The specific natural products of the primary and secondary metabolites are
very important for chemists.
7. Alkaloids are a group of naturally occuring chemical compound.
8. Nicotine is the most abundant of the volatile alkaloids in the tobacco leaf.
Molecular Formula : C10H14N2 3
INTRODUCTION

4. MEDICO ETHNO BOTANICAL SURVEY
OF Nicotiana tobaccum
Kingdom Plantae
(unranked) Angiosperms
(unranked) Eudicots
(unranked) Asterids
Order Solanales
Family Solanaceae
Genus Nicotiana
Species N. tabacum

5. Medicinal uses of tobacco
Medicinal
uses of
tobacco
Toothaches
Insect
Repellent
Rattle Snake and
Insect Bites
COLD
Tooth Paste
Anticancer
Agent
Skin rashes
Resist
Infection

6. • To extract the phytochemicals present in Nicotina tobaccum plant
using N-Hexane, Chloroform and ethyl acetate solvent.
• To perform the Phytochemicals screening of all three extracts of
Nicotina tobaccum plant.
• To determine the physiochemical parameters such as Ash values,
Fluorescence Analysis and the Qualitative determination of
Phytochemical such as Alkaloids, Ascorbic Acid.
• To study the Antibacterial and Antioxidant Properties of three
solvent extract of Nicotina Tobaccum .
• To anlayse the Phytochemical constituents using analytical
techniques:GC-MS and HPTLC fingerprinting.
• Characterization of Plant extract using IR spectroscopy, HRMS
spectroscopy they use in determination of functional groups, and
mass and Ionization state.
6
AIMS AND OBJECTIVES

7. EXPERIMENTAL SECTION
7

8. EXPERIMENTAL SECTION
drywash
Add solvent
STEP-1
HRMS
FT-IR
Spectral Studies
Fluorescence, Quantitative
determination, TLC, GC-MS.
Phytochemical screening test
Analysis
STEP-2
Biological Activity
Antioxidant
Antibacterial
STEP-3

9. RESULTS AND DICUSSION
1. Phytochemical screening tests.
2. Fluorescence Analysis
3. Estimation of Alkaloids, Ascorbic
acid, Phenolics
4. TLC Analysis
5. Antibacterial Activity
6. Antioxidant Activity
7. HPTLC Analysis
8. GC-MS Analysis
9. FT-IR Analysis
10. High Resolution–Mass Spectroscopy
9

10. 1. Phytochemical Screening:
Sl.No Phytochemicals N-Hexane Extract Ethyl Acetate Extract Chloroform Extract
1 Alkaloid + + +
2 Amino acid - - -
3 Anthraquinones - - -
4 Catechin - - -
5 Fixed oil - - -
6 Flavonoid - - -
7 Glycoside - - -
8 Phenols + + +
9 Quinone + + +
10 Reducing sugars - - -
11 Saponin + + +
12 Steroids - + +
13 Tannin + + +
14 Terpenoids - - -
15 Xanthoprotein - - -
10

11. Sl.No Particulars of treatment
Colour Appearance
Visible light UV-Light
.1 Powder as such Brown Brown
2 Powder +1N NaOH Brown Green
3 Powder+1N NaOH(aq) Brown Green
4 Powder + HCL (1:1) Brown Green
5 Powder + HNO3 (1:1) Brown Green
6 Powder + H2SO4 (1:1) Brown Dark
2. Fluorescent Analysis of the powder of Nicotiana tobaccum.
.
Figure. 1 Flourescence analysis of samples under UV light (365 nm)
(NaOH, HCL , H2SO4, HNO3, CHCl3, H2O)
11

12. 3.Quantitative Parameters
Moisture content - 3.9%
Total Ash - 39.6%
Acid soluble Ash - 44%
Water-soluble Ash - 69%
Qualitative Determination of Mineral Constituents:
12
6. Thin layer chromatography analysis

13. 1. Estimation of Alkaloids by Singh and Archana Sahu Method.
0
0.5
1
1.5
2
2.5
3
3.5
1 2 3 4 5
Adsorption
Concentration
13
0
0.5
1
1.5
2
2.5
3
3.5
1 2 3 4 5
Adsorption
Concentration
2.Estimation of Phenol
3.Estimation of Ascorbic Acid(VitaminC)
by 2.4-DNPH Method.

14. HPTLC Analysis
The HPTLC analysis of N-Hexane, Ethyl Acetate, Chloroform extract shows the presence of seven different phytochemicals in extract.
Figure. 2. HPTLC analysis of N-hexane, ethyl acetate and chloroform extract.
14

15. 15

16. GC-MS Analysis:
The results from GC-MS analysis leads to the identification of number of pharmacologically important compounds from the GC fractions of the
N-Hexane, Ethylacetate and Chloroform extract of Nicotina Tobaccum. The constituents of the plant extract were identified by comparing of
the mass fragmentation
Figure. 5.8. GC-MS analysis
16

17. Infra Red Spectroscopy Analysis
17

18. High Resolution Mass Spectrometers (HRMS) Analysis
Fig.4 HRMS spectrum of N-Hexane Solvent extract of Nicotina Tobaccum and the proposed
fragmentation paths.
18

19. 0
2
4
6
8
10
12
14
16
18
1 2 3 4
Series1
Series2
Absorbance
Antioxidant activity: ferric reducing power method
Ferric Reducing Antioxidant Power
Results
Absorbance at 700 nm
•Series 1.indicate-plant extract, series 2.indicate –standard.
19
0
0.01
0.02
0.03
0.04
0.05
0.06
0.07
0.08
0.09
1 2 3 4
Plant Extract (MD)
Ascorbic Acid (Standard)
Modified ferric iron reducing antioxidant power assay (Modified FRAP assay)

20. Antibacterial activity of N-Hexane, Ethylacetate, Chloroform extracts:
Antibacterial activity of aqueous leaf extract of Nicotiana tobaccum against five human pathogens as tested
by paper disc method.
Bacterial strains Zone of Inhibition in mm
Hexane Ethyl acetate Chloroform standard
1. B.Sabtilis 5 mm 6 mm 6 mm 10 mm
2. S.epidermis 6 mm 6 mm 6 mm 10 mm
3. E.Aerogens 7 mm 5 mm 6 mm 10 mm
4. Seudomonas aeruginosa 6 mm 5 mm 5 mm 10 mm
5. Klebsiella aerogenus. 6 mm 5 mm 6 mm 10 mm
20

21. SUMMARY AND CONCLUSION
• The work carried out on Nicotina Tobaccum, showed the presence of various
Phytochemical constituents and bioactive compounds in all the extracts in
varying range and quantity.
• The quality and quantity of phytochemicals are based on the selection of
solvent system.
• The biological activities are attributed to Phytochemical constituents of
Nicotina Tobaccum and the fluorescent analysis of powdered drug can play a
remarkable role in the determination of quality and purity of the raw drug.
• These types of studies have vital role because of the commercial and research
interest.
• The various functional groups, detected from FT-IR analysis of N-Hexane,
Chloroform and EA extracts, are responsible for the medicinal properties of
plant. The compounds identified by GC-MS in N-Hexane extract of Nicotina
Tobaccum are medicinally valuable and possess a wide variety of
pharmacological applications.
• Further isolation and testing of phytoconstituents for their individual
biological activity will undoubtedly bring promising results in the discovery
and development of novel drugs. 21

22. References
• 1..Duangsri P, Juntarapun K, and Satirapipathkul C: The tobacco leaf extract and
antibacterial activity in textile. International Conference: Textiles & Fashion
2012; 3-4.
• 2.Fransworth NR, Akerele O, Bingel AS, Soejarto DD and Guo: Medicinal plants in
therapy. Bulletin of the World Health Organization 1985; 63:965–981.
• 3.Sharma Y, Dua D and Srivastva SN: Comparative study of different parts of
Azadirachta indica (neem) plant on the basis of anti-bacterial activity,
phytochemical screening and its effect on rat PC–12 (Pheochromocytoma) cell
line. International Journal of Biotechnology and allied fields 2014; 2 (7):144 –
154.
• 4.Bakht J, Tayyab M, Ali H, Islam A and Shafi M: Effect of different solvent
extracted samples of Allium sativum on bacteria and fungi. African Journal of
Biotechnology 2011; 10:5910–5915.
• 5.Nascimento GG, Locatelli J, Freitas PC and Silva GL: Antibacterial activity of
plant extracts and phytochemicals on antibiotic resistant bacteria. Brazilian
Journal of Microbiology 2000; 31:247-256.
22

23. ACKNOWLEDGEMENTS
.
My sincere thanks to my guide Dr. M.F.Valan for taking special interest
in giving me valuable suggestions, encouragement and generous assistance
throughout this project.
I am grateful to Rev.Father Dr.JohanPragasm,SJ,LIFE,Director
Loyola College for providing all the necessary facilities.
I extend my thanks to HOD, Dr.M.GeorgeJohnson & Department of
chemistry all faculty members.
23

24. 24