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<strong>HPLC</strong> <strong>Column</strong> <strong>Selection</strong> <strong>Guide</strong>linespH and Mobile PhaseThe choice of mobile phase for a reversed-phase system starts with selecting the organicmodifier. Selectivity differences and sample retention will vary significantly among mobilephases containing acetonitrile, methanol, and tetrahydrofuran (THF). Sample solubility is likelyto differ in such solvents and dictate use of a specific solvent or solvents. UV detection atcertain wavelengths is not possible with certain modifiers (e.g., methanol at 200 nm).Both pH and ionic strength of the aqueous portion of mobile phases are important parametersin developing rugged methods that are not sensitive to small variations in conditions. Withionic compounds, retention of typical species shows significant changes with pH. It is veryimportant to control pH in such reversed-phase systems to stabilize retention and bandspacing. A pH set between 2 and 4 generally provides the most stable conditions for retentionvs. small changes in pH, and this pH is recommended for starting method development formost samples, including basic compounds and typical weak acids.<strong>HPLC</strong> <strong>Column</strong>StationaryPhase<strong>Column</strong>DimensionsType ofSurfacePore SizeParticleSizeLengthInnerDiameterChemical PropertiesChemical Lifetime/SelectivityRetention FactorPhysical PropertiesEfficiencySensitivitySpeed10 Order online at www.agilent.com/chem/store
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