Practice question

Ion exchange chromatography exploits differences in net electric charge of analytes. Stationary phase functionalized with charged groups interacts electrostatically with oppositely charged solutes. Elution is controlled by pH influencing ionization and ionic strength providing competing counter-ions. Proteins with different pI values exhibit distinct charge at a given pH, causing differential affinity to matrix. Hydrophobicity governs reversed-phase and HIC, mass governs size exclusion, color is irrelevant. Selectivity can be modulated by gradient elution. This mechanism is central to purification of amino acids, nucleotides, and proteins where charge variants require high-resolution separation under non-denaturing conditions.