Coumarins 121007101349 Phpapp02

8/13/2019 Coumarins 121007101349 Phpapp02

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GURU JAMBHESHWAR

UNIVERSITY OF SCIENCE &

TECHNOLOGYHISAR

COUMARINS

Submitted by

shiva(11124002)M.Pharm 2ndsem

(pharmacognosy)


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Extraction,isolationand

characterization of

coumarins


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CONTENT

Defination of coumarin

Classification

Extraction methods

Isolation

Identification methods

Structure elucidation


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Coumarin is 5,6-benzo-2-pyrone

derivative.

O O


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The name coumarin is derived

from the carribbean word

coumarou for the tonka treefrom which coumarin with the

characteristics aroma of new-

mown hay was isolated.


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Unsubstituted coumarin is

widespread in nature in free form or

as glycoside.

coumarins along with its derivativeoccurs abundantly in plant families

such as

umbelliferae,rutaceae,leguminacea,

orchidaceae,asteraceae,guttiferae,

thymelaceae and solanaceae.


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Coumarins play role in plant

protection and are biosynthesized

de novo in stress conditions as

phytoalexins.

In low concentration they showsynergistic activity with plant growth

promoting.


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Biological activity of coumarin is

diverse and depend on their

chemical structure.

Oldest medicine from this group

is dicoumarol ,isolated fromMelilotusalba linn. have

antithrombotic and

anticoagulative properties.


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Solubility of coumarins

depend upon phenolic

hydroxyl group and theglycosidic bond.


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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)


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Coumarins are classified

based on substitution inbenzene and pyrone

rings.


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--substituents both in benzene ring

and pyrone ring

2--furanocoumarins with

substituents on benzene nucleus orpyrone ring

3--pyranocoumarins withsubstituents on benzene and pyron

rings


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SIMPLE COUMARINS

OOH O

OOH

OH

O

AESCULIN

UMBELLIFERONE


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FUROANOCOUMARINS

Furane ring is condensed withcoumarin structure.

--Psoralen type--Angelicin type


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ANGELICIN

O O O


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PSORALEN

O

O O


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FURANOCOUMARINS

Solubility-- lipid soluble

Isolation-- by extraction with ether/light

petroleum

SeparationTLC

Adsorbentsilica gel

Solventether-benzene(1:1)

Development time1 to 2hours

Detectionunder UV light


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PYRANOCOUMARIN(linear)

O O O

CH3

CH3

XANTHYLETIN


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PYRANOCOUMARINS(angul

ar)

O OO

OH

CH3

CH3

LOMATIN


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COUMARIN GLYCOSIDEAcsulin- bark ofAesculus

hippocastanumfamily-Hippocastanaceae

Cichorin-flowers of Cichorium intybuslinn

family-Compositae

Daphnin-bark of Daphne mezeriumfamily-Thymelaceae

Fraxin-bark of Fraxinus excelsir linn.

family-Oleaceae


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FURANOCOUMARIN

GLYCOSIDE Khellol-seeds of Eranthis hyemalis linn

family-Ranunculaceae

Visnagin-seeds ofAmmi visnaga

family-Umbelliferae

Psoralea-dried ripe fruit of Psoralea

corylifolia linn

family-Leguminaceae


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CLASSICAL EXTRACTION

METHODS

continous hot percolation

maceration

percolation

ultrasonic-aided extraction


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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

Continous hot percolation n


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supercritical fluid extraction

ultrasonification aided extraction

microwave-assisted solvent

extraction

pressurised liquid extractionmedium pressure solid liquid

extraction

accelerated solvent extraction


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SUPERCRITICAL FLUID

EXTRACTION

Principle- supercritical fluid is applied asan extractant

Advantages- high process speed

lack of organic solventpossibility of coupling with

Other methods like GC,HPLC

Used for separation of furanocoumarinsfromAngelica archangelica.


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isolation

Based on lactone type of coumarinstructure

PROCEDUREalcoholic solution ofpottassium hydroxide crashes the

lactone ring in coumarin result in

coumaric acid

After acidification these acid cyclize to

coumarin again


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NEW METHODS OF

ISOLATION Sublimation and fractionating

distillation in high vaccum

Crystallization from organic solvents

Distillation with water vapours


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GENERAL CHEMICAL TEST

3 volume propylene glycol5 volume acetic acid43 volume water and shake

blue fluorescence under UV light

NaOH solution yellow fluorescence

under UV light

DRUG

DRUG


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TLC of coumarins

Adsorbentsilica,florosil,polyamide,alumina

(normal phase)

Silanized silica(reverse phase)

Eluent---medium polar solvent like

dichloromethane(normal phase)aqueous solvent(reverse

phase)


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TLC of furanocoumarins and

pyranocoumarins

Adsorbentsilica, florisil, polyamide,

alumina

Eluentweak polar solvent:

petrol+diethyl ethertoluene+ethyl acetate

n-hexane+ethyl acetate


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Rf VALUE OF

FURANOCOUMARIN

FURANOCOUMARI

NS

ETHER:BENZENE CHLOROFORM

Bergapten 100 100

Isobergapten 112 167

Pimpinellin 108 86

Isopinellin 97 43

sphondin 92 90


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2-D TLC

Adsorbent-diol silica(polar bonded)

1stdirection eluent-10% methanol inwater

2nddirection eluent-100% diisopropylether

PRINCIPLE-coumarins are identified bycomparing retardation factors in bothdirections.


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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 isapplied as band on silanized silica

layer eluted with methanol:water(6:4).


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UV-SPECTROSCOPY

-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


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SPECTRAL DATA OF

HYDROXYCOUMARINSCOUMARIN EtOH max 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


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STRUCTURE ELUCIDATION

IR Spectroscopy

NMR

Mass spectoscopy


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INFRARED

SPECTROSCOPY Stretching frequency -1700-

1750cm(c=o)

Skeletal vibrations-1600-1660cm(c=c)

Stretching frequency-1500cm(aromatic

ring)


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H-NMR SPECTROSCOPY

NMR spectra of coumarins H-3 and H-4protons exhibits characterstics chemical shiftwhich distinguish different coumarin

Upfield shift of 0.17 ppm of H-3 Proton in 7-oxygenated coumarins as compared withcoumarins is due to electron release resultingin electron density at C-3

An oxygen at C-5 shift the resonance of H-4downfield by 0-3 ppm due to peri effect


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H NMR chemical shift of coumarins

Coumarinderivatives H-

(ppm) H-

(ppm) (Hz)

Coumarins

5-oxygenated

7-oxygenated

8-substituted5,7-

disubstituted

6-substituted

Angular

furanocoumar

ins

6.1-6.4

6.1-6.4

6.2-6.5

6.6-6.96.7

6.7-7.2

7.5-7.7

7.5-7.9

7.9-8.2

7.6-7.8

7.1-7.56-7

7.5-7.7

6.7-7.2

9.5

9.5

9.5

8.52.5

2.5

2.5


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MASS SPECTROMETRY

Coumarin on electron impact gives astrong molecular ion peak (M) at m/e146 (76%) and a base peak at m/z118(100%) by the loss of 28 mass unitsequivalent to carbon monoxide

7-hydroxy coumarin show a strong Mionat m/e 162(80%) and base peak at m/z134 due to loss of carbon monoxide


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MASS FRAGMENTATION

PATTERN OF COUMARIN

AND FURANOCOUMARINS.


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coumarinsO

O

O

m/e 146(76%)

m/e 118(100%)


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7 hydroxycoumarins

O OOH

OH O

m/e 162(80%)

m/e 134(100%)


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REFERENCES

Harborne J.B,Phenylpropanoid,A guideto modern techniques of plant analysis2nd,champan and hall,New york 44-47

Kar Ashutosh ,coumarin

glycoside,pharmacognosy andpharmacobiotechnology,new age Ltd.New delhi 512-515

Kowalska teresa,application of tlc in

analysis of coumarin,TLC inphytochemistry,CRC press, London 78-84

Bhatt S.V chemistry of natural productsnorosa pulisher 412-416

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Coumarins 121007101349 Phpapp02