this ppt. describe the coumarin present in herbal drugs and plant sources.

1. GURU JAMBHESHWAR
UNIVERSITY OF SCIENCE &
TECHNOLOGY
HISAR
― COUMARINS‖
Submitted by
shiva(11124002)
M.Pharm 2nd sem
(pharmacognosy)

2. Extraction,isolation
and
characterization of
coumarins

3. CONTENT
 Defination of coumarin
 Classification
 Extraction methods
 Isolation
 Identification methods
 Structure elucidation

4. Coumarin is 5,6-benzo-2-pyrone
derivative.
O O

5. The name coumarin is derived
from the carribbean word
―coumarou‖ for the tonka tree
from which coumarin with the
characteristics aroma of new-
mown hay was isolated.

6. Unsubstituted coumarin is
widespread in nature in free form or
as glycoside.
coumarins along with its derivative
occurs abundantly in plant families
such as
umbelliferae,rutaceae,leguminaceae
,
orchidaceae,asteraceae,guttiferae,
thymelaceae and solanaceae.

7. Coumarins play role in plant
protection and are biosynthesized
―de novo‖ in stress conditions as
phytoalexins.
In low concentration they show
synergistic activity with plant growth
promoting.

8. Biological activity of coumarin is
diverse and depend on their
chemical structure.
Oldest medicine from this group
is dicoumarol ,isolated from
Melilotus alba linn. have
antithrombotic and
anticoagulative properties.

9. Solubility of coumarins
depend upon phenolic
hydroxyl group and the
glycosidic bond.

10. Coumarins occur in plants in free
form or as glycosides.
Aglycone are soluble in
petrol,benzene,ether,chloroform,
diethylether,alcohol(nonpolar
solvent)
Glycosides are soluble in water and
alcohols(polar solvent)

11. Coumarins are classified
based on substitution in
benzene and pyrone
rings.

12. 1--simple coumarins with
substituents both in benzene ring
and pyrone ring
2--furanocoumarins with
substituents on benzene nucleus or
pyrone ring
3--pyranocoumarins with
substituents on benzene and pyrone
rings

13. SIMPLE COUMARINS
O O
HO
HO
AESCULIN
HO O O
UMBELLIFERONE

14. FUROANOCOUMARINS
Furane ring is condensed with
coumarin structure.
--Psoralen type
--Angelicin type

15. ANGELICIN
O O O

16. PSORALEN
O O O

17. FURANOCOUMARINS
 Solubility-- lipid soluble
 Isolation-- by extraction with ether/light
 petroleum
 Separation—TLC
 Adsorbent—silica gel
 Solvent—ether-benzene(1:1)
 Development time—1 to 2hours
 Detection– under UV light

18. PYRANOCOUMARIN(linear)
H3C
O O O
H3C
XANTHYLETIN

19. PYRANOCOUMARINS(angul
ar)
OH
H3C
H3C
O O O
LOMATIN

20. COUMARIN GLYCOSIDE
 Acsulin- bark of Aesculus
hippocastanum
family-Hippocastanaceae
 Cichorin-flowers of Cichorium intybus
linn
family-Compositae
 Daphnin-bark of Daphne mezerium
family-Thymelaceae
 Fraxin-bark of Fraxinus excelsir linn.
family-Oleaceae

21. FURANOCOUMARIN
GLYCOSIDE
 Khellol-seeds of Eranthis hyemalis linn
family-Ranunculaceae
 Visnagin-seeds of Ammi visnaga
family-Umbelliferae
 Psoralea-dried ripe fruit of Psoralea
corylifolia linn
family-Leguminaceae

22. CLASSICAL EXTRACTION
METHODS
continous hot percolation
maceration
percolation
ultrasonic-aided extraction

23. Continous hot percolation n
 Dried sample, ground in to small
particles and placed in a porous
cellulose thimble
 Thimble placed in extracted
chamber followed by flask
heated with solvent and a
condenser

24. MODERN TECHINQUE
supercritical fluid extraction
ultrasonification aided extraction
microwave-assisted solvent
extraction
pressurised liquid extraction
medium pressure solid liquid
extraction
accelerated solvent extraction

25. SUPERCRITICAL FLUID
EXTRACTION
Principle- supercritical fluid is applied as
an extractant
Advantages- high process speed
lack of organic solvent
possibility of coupling with
Other methods like GC,HPLC
Used for separation of furanocoumarins
from Angelica archangelica.

26. isolation
 Based on lactone type of coumarin
structure
 PROCEDURE—alcoholic solution of
pottassium hydroxide crashes the
lactone ring in coumarin result in
coumaric acid
 After acidification these acid cyclize to
coumarin again

27. NEW METHODS OF
ISOLATION
 Sublimation and fractionating
distillation in high vaccum
 Crystallization from organic solvents
 Distillation with water vapours

28. GENERAL CHEMICAL TEST
DRUG 3 volume propylene glycol
5 volume acetic acid
43 volume water and shake
blue fluorescence under UV light

DRUG
NaOH solution
 yellow fluorescence
under UV light

29. THIN LAYER
CHROMATOGRAPHY
TLC IS used for identification of
compounds presented in plant
extract by retention parameters as
well as UV spectra taken directly
from the layer by densitometry.

30. TLC of coumarins
Adsorbent –
silica,florosil,polyamide,alumina
(normal phase)
Silanized silica(reverse phase)
 Eluent---medium polar solvent like
dichloromethane(normal phase)
aqueous solvent(reverse
phase)

31. TLC of furanocoumarins and
pyranocoumarins
 Adsorbent—silica, florisil, polyamide,
alumina
 Eluent—weak polar solvent:
petrol+diethyl ether
toluene+ethyl acetate
n-hexane+ethyl acetate

32. Rf VALUE OF
FURANOCOUMARIN
FURANOCOUMARI ETHER:BENZENE CHLOROFORM
NS
Bergapten 100 100
Isobergapten 112 167
Pimpinellin 108 86
Isopinellin 97 43
sphondin 92 90

33. 2-D TLC
 Adsorbent-diol silica(polar bonded)
 1st direction eluent-10% methanol in
water
 2nd direction eluent-100% diisopropyl
ether
PRINCIPLE-coumarins are identified by
comparing retardation factors in both
directions.

34. PREPARATIVE TLC
PRINCIPLE-Components are applied in
the form of a band and
rechromatography of partly separated
fractions.
For separation of coumarins from
Heracleum sosnowskyj fruits extract is
applied as band on silanized silica
layer eluted with methanol:water(6:4).

35. UV-SPECTROSCOPY
 ὰ-pyrone-300 nm
 Unsubstituted coumarins-274nm and
311nm
 7-hydroxycoumarins-217nm and 315-
330nm
 Linear furanocoumarins-205-225nm and
240-255nm
 Angular furanocoumarins-240-255nm and
260-270nm are abssent

36. SPECTRAL DATA OF
HYDROXYCOUMARINS
COUMARIN EtOH ʎmax WATER HOAc UV light BAW
AGLYCONE
coumarin 212,274,282,312 67 76 None 92
umbelliferone 210,240,325 57 60 Bright blue 89
aesculetin 230,260,303,351 28 45 Blue 79
scopoletin 229,253,300,346 29 51 Blue violet 83
daphnetin 215,263,328 61 54 Pale yellow 81
GLYCOSIDE Water BN BAW
aesculin 224,252,293,338 56 13 Clear blue 53
scopolin 227,250,288,339 64 44 mauve 53

37. STRUCTURE ELUCIDATION
 IR Spectroscopy
 NMR
 Mass spectoscopy

38. INFRARED
SPECTROSCOPY
 Stretching frequency -1700-
1750cm⁻₁(c=o)
 Skeletal vibrations-1600-1660cm⁻₁(c=c)
 Stretching frequency-1500cm⁻₁(aromatic
ring)

39. ₁H-NMR SPECTROSCOPY
 NMR spectra of coumarins H-3 and H-4
protons exhibits characterstics chemical shift
which distinguish different coumarin
 Upfield shift of 0.17 ppm of H-3 Proton in 7-
oxygenated coumarins as compared with
coumarins is due to electron release resulting
in electron density at C-3
 An oxygen at C-5 shift the resonance of H-4
downfield by 0-3 ppm due to peri effect

40. ₁H NMR chemical shift of coumarins
Coumarin H₃-ᵹ(ppm) H₄-ᵹ(ppm) ᵹ(Hz)
derivatives
Coumarins 6.1-6.4 7.5-7.9 9.5
5-oxygenated 6.1-6.4 7.9-8.2 9.5
7-oxygenated 6.2-6.5 7.6-7.8 9.5
8-substituted 6.6-6.9 7.1-7.5 8.5
5,7- 6.7 6-7 2.5
disubstituted
6-substituted 6.7-7.2 7.5-7.7 2.5
Angular
furanocoumar 7.5-7.7 6.7-7.2 2.5
ins

41. MASS SPECTROMETRY
 Coumarin on electron impact gives a
strong molecular ion peak (M⁺) at m/e
146 (76%) and a base peak at m/z
118(100%) by the loss of 28 mass units
equivalent to carbon monoxide
 7-hydroxy coumarin show a strong M⁺ion
at m/e 162(80%) and base peak at m/z
134 due to loss of carbon monoxide

42. MASS FRAGMENTATION
PATTERN OF COUMARIN
AND FURANOCOUMARINS.

43. coumarins
O O
m/e 146(76%)
O
m/e 118(100%)

44. 7 hydroxycoumarins
HO O O
m/e 162(80%)
HO O
m/e 134(100%)

45. REFERENCES
 Harborne J.B,Phenylpropanoid,A guide
to modern techniques of plant analysis
2nd,champan and hall,New york 44-47
 Kar Ashutosh ,coumarin
glycoside,pharmacognosy and
pharmacobiotechnology,new age Ltd.
New delhi 512-515
 Kowalska teresa,application of tlc in
analysis of coumarin,TLC in
phytochemistry,CRC press, London 78-
84
 Bhatt S.V chemistry of natural products
norosa pulisher 412-416