The thermodynamics and energetics of the ion exchange of four amino acids at a synthetic ion exchange resin have been studied. Experimental work included determination of ion exchange isotherms and the use of high-sensitivity titration microcalorimetry. A rigorous thermodynamic analysis of the data is developed allowing calculation of the standard free energy, the standard enthalpy, and standard entropy of exchange, and also the differential free energy, incremental enthalpy, and incremental entropy of exchange. The results show that the relative contributions of the enthalpy and entropy to the overall free energy differ markedly for the chosen amino acids. From a knowledge of these fundamental thermodynamic properties, the solution conditions giving enhanced separation are predicted. These predictions are experimentally confirmed. The paper establishes experimental protocols and data analysis methods that allow progression from a single isotherm and a single set of calorimetric data to a selection of process conditions for enhanced selectivity. The experimental techniques and analysis procedures developed are generally applicable to ion exchange separations of biomolecules.