Agilent HPLC Column Selection Guide

HPLC Column Selection GuidelinesSmall MoleculesMW < 2000Large MoleculesMW > 20005 and 3.5 µmStandard Porous Particle Size5 µmFor conventional analyses of small and large molecules, the standard particle size is 5 µm. However, smaller particle sizes are availableand provide higher efficiency and higher resolution. They are also available in short column lengths.A 3.5 µm particle may be the preferred starting particle size because it provides scalability up to 5 µm and down to 1.8 µm without requiring method revalidation.Fast Analysis Particle SizesRapid Resolution 3.5 µmRapid Resolution HT 1.8 µmRapid Resolution HD 1.8 µmRapid Resolution 3.5 µm particles provide 60% greater efficiency than 5 µmparticles while Rapid Resolution HT 600 bar and Rapid Resolution HD 1200 bar1.8 µm particles provide 200% greater efficiency. Shorter columns can be usedto significantly reduce analysis time of small molecules.Rapid Resolution 3.5 µmPoroshell 5 µmFor peptides, Rapid Resolution 3.5 µm particles can be used withshorter columns for faster, high-resolution gradient analyses.But for proteins, Poroshell columns, with a solid core andporous shell, provide the fastest analysis.150 mm Column Length150 mmA conventional column length is 150 mm or 250 mm for high resolution with5 µm particles. This is a good place to start a typical analysis because it will providehigh resolution in a 20-30 minute analysis. For fast analysis, shorter columns(75 mm and 50 mm) with smaller particle sizes (3.5 µm) should be used.For proteins and peptide digests, 150 mm is also a good starting column length.The typical gradient time on a column of this length will be 30-60 minutes fora high-resolution analysis. For fast analysis, shorter columns (50 mm)with smaller particles (3.5 µm) should be used.Starting Column ChoicesSmall MoleculesLarge MoleculesStandard AnalysisFast AnalysisStandard AnalysisFast AnalysisEclipse Plus C184.6 x 150 mm, 5 µm959993-9024.6 x 150 mm, 3.5 µm959963-902Rapid ResolutionEclipse Plus C184.6 x 75 mm, 3.5 µm959933-902Rapid Resolution HT,600 barEclipse Plus C184.6 x 50 mm, 1.8 µm959941-902Narrow Bore RRHD,1200 bar *Eclipse Plus C182.1 x 50 mm, 1.8 µm959757-902ZORBAX 300SB-C184.6 x 150 mm, 5 µm883995-902Rapid ResolutionZORBAX 300SB-C184.6 x 50 mm, 3.5 µm865973-902Poroshell 300SB-C182.1 x 75 mm, 5 µm660750-902* First choice for use on the 1290 Infinity LC or other UHPLC instruments with 1000 barpressure limitsOrder online at www.agilent.com/chem/store7