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Ion Exchange Chromatography

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ComponentsSame as any HPLC Instrument

ReservoirsPumpInjectorColumnDetectorChromatography Software

Specific Components

Membrane Suppressor

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Ion Exchange Chromatography

Basic Separation Principle: Exchange equilibria between ions in solution and ions (of the same charge) held by surface bonded functional groups

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Ion Exchange Functional Groups

Cation ExchangeSulfonic acid – SO3

- (H+)

Carboxylic Acid

Anion ExchangePrimary Amine – NH3

+(OH-)

Tertiary Amine

– C – O-(H+) =O

– N – CH3

CH3

CH3

+

(OH-)

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Mobil Phase

Cation Exchange – H+

Anion Exchange - OH-

pH is controlled with a Buffer Solutione.g. HCO3

-/CO3--

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Stationary Phase – Ion Exchange Resin

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Stationary Phase – Solid Support “Backbone”

CH CH2 CH CH2

CH CH2

styrene Divinylbenzene (DVB)

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Polystyrene Divinylbenzene

R R

RR R

CH CH2 CH CH2CH CH2

CH CH2 CH CH2CH CH2 CH CH2

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1. Cellulose (the cellulose, derived from wood or cotton, is lightly derivatized with ionic groups to form the ion exchanger).

Anion exchange

Cation exchange

2. Gel-type ion exchangers: combine the separation properties of gel filtration with those of ion exchange. Because of their high degree of substitution of charged groups, which results from their porous structures, these gels have a higher loading capacity than do cellulosic ion exchangers.

Stationary Phase – Ion Exchange Resin

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Separation of Ca2+ Mg2+ Sr2+ and Ba2+

Exchange equilibrium at the head of the column:2(R-SO3-H) + M2+ = (R-SO3)2-M + 2H+

solid solution solid solution

Separtion/Elution begins when acid solution is added

Equilibrium shifts to the leftCation goes into the mobil phase (momentarily)Cation desorbs/re-adsorbs many times

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The time the cation spends in each phase is governed by its unique value of Kexchange

Rearranging gives:

Separation of Ca2+ Mg2+ Sr2+ and Ba2+

aq2

solid2-

3

aq2

solid2

2-

3reaction exchange

][M ]H[RSO][H M)[(RSOK

++

++

=]

aq2

solid2-

3ex

aq2

solid2

2-

3

][H]H[RSO K

][MM)[(RSO

+

+

+

+

=]

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Separation of Ca2+ Mg2+ Sr2+ and Ba2+

The terms on the r.h.s. are constant when:

[H+]aq > [M+2]aq

[RSO3-H]s > [(RSO3

-)2M2+]s

i.e. [acid] > [cation] and binding sites are not saturated

aq2

solid2-

3ex

aq2

solid2

2-

3

][H]H[RSO K

][MM)[(RSO

+

+

+

+

=]

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Separation of Ca2+ Mg2+ Sr2+ and Ba2+

Normal Chromatography Separation Principles Apply:

M

SD

aq2

solid2

2-

3

CC K

][MM)[(RSO

==+

+ ]

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Ion Exchange Chromatography –General Characteristics

In general, ion exchangers favor binding of ions with:

↑ charge3+ > 2+ > 1+

↓ hydrated radiusH+ > Li+ > Na+ > K+ (p. 174)H+ bound least strongly b/c hydrated radius is highest

↑ polarizability

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ExampleChromatograms

Cation ExchangeAnion Enchange

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Example Chromatograms

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Illustration of Ion Exchange Chromatography