Chromatography Forum

Hi everyone,

Basically I can't think why the pressure on a HPLC pump will vary when the isocratic mobile phase is changed.

I know that solvent composition is a maintained constant throughout the isocratic elution, but it sounds too easy to say that pump pressure would correlate with a change in isocratic mobile phase i.e. increase in mobile phase would increase the pressure on the pump?

Unless i'm getting confused.

Any help would be great!

You are getting yourself confused (don't worry, happens to all of us now and then!)

The pressure at the pump is determined by two things:
- the flow rate (pressure is directly proportional to flow)
- the flow resistance of the system.

The flow resistance is (usually) dominated by the column, and will depend on:
- the dimensions of the column (directly proportional to length, and inversely proportional to the square of the column internal diameter)
- the particle size of the packing material (inversely proportional to the square of the particle size)
- the viscosity of the liquid (directly proportional to viscosity). Viscosity depends on liquid composition and temperature.
Aside from that, things like plugged filters, or kinked or very narrow tubing can contribute to flow resistance.

"Isocratic" is just a fancy word that says mobile phase composition and flow stay constant during any given run. So pressure should stay constant during an isocratic run.

Are you changing the composition of the mobile phase? If so then you could be changing the viscosity which would give a different pressure all other things being constant. Even if you make fresh mobile phase, if it is a buffer and the concentration or pH differs slightly then the pressure could change slightly.

Also is your column in a heated/cooled compartment or open to ambient air? Columns that are open to ambient air will cause pressure changes with changing room temperature.