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Ion Exchange Chromatography

Ion Exchange Chromatography. Uses displacement of an ion from the stationary phase by a solute ion K is dependent on relative affinity of the analyte for the surface over the solvent ions. Principles Small K means low affinity – so low RT

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Ion Exchange Chromatography

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  1. Ion Exchange Chromatography Uses displacement of an ion from the stationary phase by a solute ion K is dependent on relative affinity of the analyte for the surface over the solvent ions Principles • Small K means low affinity – so low RT • K is often quoted vs. a common reference ion (e.g. H+) • K affected by: • Charge of ion:  charge gives  K • Size of hydrated ion

  2. Cation Exchange Chromatography Use organic acid functional groups as ionic species “Strong acid” type uses sulphonic acid (RSO3H) (more common) “Weak acid” type use carboxylic acids (RCOOH) xRSO3H + Mx+ (RSO3-)xMx+ + xH+ Stationary Mobile Stationary Mobile Ion Series For a strong acid column, (e.g. RSO3H) size of K : Ag+ > Cs+ > Rb+ > K+ > NH4+ > Na+ > H+ > Li+ and Ba2+ > Pb2+ > Sr2+ > Ca2+ > Ni2+ > Cd2+ > Cu2+ > Co2+ > Zn2+ > Mg2+

  3. Anion Exchange Chromatography Use amine groups as ionic species “Strong base” type uses quaternary amines (e.g. RN(CH3)3+OH-) “Weak base” type uses secondary or tertiary amines xRN(CH3)3+OH- + Ax- (RN(CH3)3)x+Ax- + xOH- Stationary Mobile Stationary Mobile Ion Series For a strong base column, affinity series for K is usually : SO42- > Cr2O42- > I- > NO3- > Br - > Cl- > HCO2- > CH3CO2- > OH- > F-

  4. Ion Exchange Chromatography(optimisation) Stationary phase: “Strong” or “weak”, different pKa’s and pKb’s of ionic stationary phase groups give different strengths Mobile phase • pH: can affect both a) the proportion of protonated stationary phase sites, b) the proportion of ionised analyte, but b) is more influential so  in mobile phase pH gives: •  in retention time for cation exchange •  in retention time for anion exchange • Ionic Strength: (concentration of the mobile phase), mostly increases selectivity between ions of different charge • Buffer salt: i.e. the relative position of the competing counter ion in the series for K • Organic modifier: resin supports have a non-ionic component so reverse-phase interactions can occur. Affected by adding an organic solvent • Temperature: affects buffer solubility, viscosity and mass transfer kinetics

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