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Column Chromatography 10m - RS

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14 views5 pages

Column Chromatography 10m - RS

Uploaded by

iamidiot666462
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© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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3 colmn chromatography

wDeftnitìon ouloe
colunn chronatography is a tye of chromatograpy
In which Lhe statiorary Phale is Pocked in the
column (Glas y Metal co lurnn) and t is usd
For the separation crd Rrification cf Substonce
WPinciple
* The pânciple Involved is Adaorption chonatogidphy .
Biding of compame on a statiorayy phase is
calledà asorption.
Components with higher affinity for stoti onary phae
move souo and touer afiaty wil move foat.
* Analyte
The sarnple to be separcted is caled Aralle.
(Components)
Stationayy phaie
which nteract
’IE is a stable and fìxed. part
oith the moleleg to be separated.
’ Eg: silica Gre|.

* Mobile phse
Tt is Ahe movable par t phich helps Lhe
Analyte to Interact oith statiorars plaje
>g:, Solent that Tun throug h the colron is a
mobile @hoge.
() Expeinental Techlque

’ Tacking of column
*A cylindrical glass Pod is used a column and tE is
held in a Ver bCal Position .
solld Adsorbent atenals ike sla gel or Alurnina
(statio naay phaJe) is PAcked into the colurnn to
Create a Uniform bed.
* Packing cf column is cone by too Methods
-sPry FoCking The Dry Adsorbent is added in the
coumn, and then solvent is gently ahod)
et parking CTho stati onay phaje is mixed oE
Sovent to form a slurry. which is ten pouYec
into the column)
’, APplcation of Sanple
snall
The mixture to be separated is dissolved in a
Volune o solventA.
the top of the
* The Sample is Carefully appled topiretie
Fucked column uoth the help of .

choice f solvert
The solvent used to cary the copourds th ough the
Called the eluent (Mobile phaye) Colunn is

There ae many solents use in the coluna chona tarapky


and thecfee depends on polariy and Soubi lity.
Choice
Methand
Cemmon eluent incudo, - Hexane, Etyl acetate
dnd water.
4 Development oF the column

conponevnts ef the arple oe sepa vats by


The
4he continuou pasoge f the mobile phase throUg h
Lhe column.
Flew cf solvents is continue until the Mixhi
* The
complelty sepoarated into the comporent
is

Mobile phase Celuent)


Sample
Filter paer disc/sand
stbroger
Iterackion stationavy phase Csilica )
Weakor Glass od colurmn
Interaction
cotton/
ghas wbol
Column -Fracions collection
chroratog Kaply
*Three conmon elution mnethods
OSimple or lso (ratic Elton
Single solvent is Use a mobile phaso
)SLepoi se er bath ELution
Gradually changing solwent ding chromaigraphy,
particularly wueful when the mixture ho diferent
polaritieg
(ijGyadient Eluthon
clar ing uith a legs polar solvent arot vaddlly
Increairg polority of the solvent
Co lection of Froactiorg
Dfferent compouds are collectsd separa tely ussally
in test tubes or Vials )
De tehon and Identi fiabon
compoents detecton is easier and i bajed
t Coloured
on Visual observation

Tf Lhe, compours o e soated from Lolumn are


colcuré ss, Ulba vìolet rMetheds are generally uge
The columns are direcy Eamined urder ÚV lamps.
y Thin Layer chrematagraphy Small Fractios of the
Eluent ore collected and eoch rattion is araly sed
by T L C . 2
* Other techniqeg Fluorescent SpechoscoPy, Padio
active emission, ete
the
Chromatogran - Guaphical sepreßentotion of
obAoined fiorn chronatography EZperiment.
Pesu
conyonents
TE displays the seraotion of ditfeet
hromatography
in mìxure a they ehte fiom the
column ONer Lime.

9Types
Ion Excharge choratogiqphy
* Affinity chrorgtogiaphy
FlLaaion Chromatography
* Grel
Liquid chrornatograhy
* Gias chrgtoqaphy )
HPLC (High pressure Lqwda
FPLC (Fast Protein Liquúd chronatography.
(APplication
Separatien of tixture oF ornonents
Cpigmeats, ipida, sugars)
Puification Removal cf lpurifies
-Protein Turificationbhei
Aucleic Acid purificaion
- Forensic Applhication To Detect and asarih,
drugs, isond and other ox0C Substanca! ia
Biolegical Sargplei CBlod Vine,ek)
Food Indttry - in Gc, byty separating and
analysing additves, Presernativel,etc,

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