ion exchange chromatography

simple form of IEC

1. Ion exchange chromatography
Presented by
Liji jesu

2. INTRODUCTION
•ION EXCHANGE CHROMATOGRAPHY
is a process by which mixture of similar
charge ions can be separated by using ion
exchange resins.

3. principle
•The reversible exchange of ions in
solution with ions electro statically bound
to inert support medium.

4. Factors in ion exchange
•Electrostatic force of attraction
•Relative charge
•Radius of hydrated ions
•Degree of non bounding interactions

6. Ion exchangers
Columns are packed with ion exchangers
•Anion exchangers
•Cation exchangers

8. Counter ions
•Ions bounds electro statically to
exchanger.

9. ELUTING X+
Increasing conc.
Of y+
Increasing ph of
solvent
ie converting x+
to uncharged sps
Y+ conc to elute
x+ depend on
quantity of x+
charge

10. Macromolecules
Ptns and nucleic acid which can posses
+ve
And –ve charge
It can bind ,anion and cation exchangers
Make them +ve by increasing ph
make –ve by reducing ph

11. Types of ion exchange resins
•Poly styerne
•Cellulose
Properties of resins
•Ion accessibility
•Chemical stability
•Mechanical stability

12. type nature
strong cation Salphonated polystyrene
weak cation Sulphoproponyl cellulose
Condensed acrylic acid
Carboxy methylcellulose
Strong cation Dimethyl
Quadatory amino cellulose
weak anion Dithylaminoethyl cellulose
Dithylaminoethyl agarose

13. Pptn of exchange medium
•Swelling of medium
•Removal of very small particles
•Exchanger lies to equilibrium

14. Use of buffer
•To maintain ph of column.
•Anionic IEC
•Cationnic IEC

15. BUFFER PH range
Ammonium acetate 4-6

Ammonium formate

3-5
Pyridium acetate 4-6
ammonium carbonate 8-10

16. procedure
•Ptn mixture transferred into low ionic
strength .(mb)
•absorbent is packed into a column the
column is pre equilibrated with the buffer
of identical ph and similar ionic strength
as protein mixture (prefr the same buffer
as ptn mixture)
•Ptn mixture is applied into column .ptns
charged oppositely to IE media are
temporarily retained in column .

17. •All other ptns simply pass through the
column and are collected during this
step..
•Retained ptns are eluted from the column
by applying a modified buffer ,elution is
most commonly achieved by gradually
increasing their ionic strength of buffer via
salt gradient ,and ptn are eluted in order
of increasing their net charges, is specific
cases the elution can be accomplished.

18. •a)A ph change
•B)Affinity methods

20. APPLICATIONS
OF IE CHROMATOGRAPHY
•Use for amino acid analysis
•To determine the composition of nucleic
acid.
•Uses for water purification
•Separations of vitamins ,biological amines
and bases
•Separation of ultra pure metals

22. Thank you