Column Evaluation Method Development

Six methods for evaluating reversed phase (RP) column and six methods forevaluating strong anion exchange (SAX) column are compared. Advantages anddisadvantages of these methods are obtained by compared system stability,dead time,theoretical plates,asymmetry and analysis time, which are benefit to producer and user.New methods for evaluating RP and SAX are developed based on the comparisonresearch. To the RP column, the recommended composition of the mobile phase is 90%methanol and 10% buffer, where the buffer composition is 20 mmol/L potassiumphosphate adjusted to pH 7.0±0.1, and the sample is uracil, toluene and amitriptyline.Uracil is used as an indicator of the void volume. Toluene is a useful marker forcalculation of column efficiency (theoretical plates, N) and hydrophobility of the packing.Amitriptyline is a basic (pKa=9.4) pharmaceutical drug (antidepressant) used for columncharacterization. To the SAX column, the recommended composition of the mobile phaseis 50 mmol/L potassium phosphate, and the sample includes adenosine,CMP,AMP,UMPand GMP. Adenosine is used as an indicator of the void volume. GMP is a useful markerfor calculation of column efficiency (theoretical plates, N).