In a separation of a basic drug analyte from its impurity using strong cation exchange chromatography (HPLC), using 150 mM of ammonium dihydrogen phosphate at pH 3 and flow of 1.4 ml/min, the resolution is very low.
How to get good separation, what are the variable to vary in the mobile phase.
by modelling: you need two gradients from low to high salt concentration, (f.e. from 10 mMol/L to 200 mMol/L or even higher) at 30 and 60°C, with the same start- and endconcentrations, inject the same sample and the same amount. These are 4 experiments at constant pH. Than enter them into DryLab (we can do this for you), which will create ca. 10000 different experiments and you can find the best one with a mousclick. See more info at www.DryLab.com
You could change pH, using another buffer, or mix of two solvents as mobile phase. It depends of drug structure and pK of drug and impurities.
Yes, but pls use gradient elution. Don't waste your time with isocratic experiments.
thanks a lot for your suggestions Imre. yes i agree with you about the gradient. but why to choose different temperatures.
We do not have dry lab.
because the separation might be different at different temperatures and you can find a better separation.
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