Chromatography Forum

I'm trying to get a method working with an Agilent Hi-Plex ion exclusion column, which is super-fiddly because it seems like you need to treat it with kid gloves for it just to survive, let alone work properly. I'm trying to see oxalic acid using a 1200 HPLC and 6410 triple quad, for context.

These columns apparently need a slow build up in flow rate (0.1>0.2 mL/min) and organic phase (0>20% MeCN), so I've made up a 'startup' method that will ramp the flow rate and mobile phase comp. over about 10 minutes.

The problem is that when MassHunter Acquisition switches to the 'run' method for analysis (which is isocratic and matches the final conditions of the 'startup' method), there's a split second when it changes when MassHunter reverts to the initial conditions (e.g. 0.1 mL/min and 100% buffer A) of the 'startup method' before switching to the 'run' method. I'm concerned that this abrupt shift in composition and flow rate might slightly damage this expensive column.

So the question is - is there a way to enable MassHunter to just keep going with the final conditions of the method and not revert to the initial conditions at the end of a run? I feel like there's something obvious I'm missing here...

Does your software have a spot for "post run conditions"? Some versions have this and may be helpful.

Does it switch back if you load the isocratic method manually?

If not then the option would be to switch to the run method manually before starting the acquisition. That does mean you have to baby sit it until the run starts and can't just start the equilibration prior to walking out the door, but might save the column.

Knowing Agilent/MassHunter quirks ... can we be certain that it isn't just a graphical software artifact that it says it reverts back to the starting conditions momentarily only on screen whereas the actual pump conditions don't revert back? Do you see a pressure change to confirm if it actually did drop back down to 0.1 mL/min? If it happens fast enough that a pressure change doesn't happen then I wouldn't panic.

I do second James_Ball's idea though about equilibrating it manually instead of with a method, I suppose it depends on if your beginning column equilibration needs 30 min or several hours.

If you were doing this manually, you wouldn't do it as a totally smooth gradual increase, you'd do it stepwise, increasing the flow and adjusting the percentage a little at a time. How many steps would you take, to do it manually?

If it's not a huge number, say 5 steps, you could set up 5 methods running those percentages and flow-rates, and prepare yourself a sample-list that begins with those 5 methods run sequentially. It won't matter that it goes back to the starting conditions in each of these methods, because they have only one flow and percentage; they're simply mini-isocratic methods. But each one is a bit higher flow than the one before, so by the time you've run all 5, you've gone stepwise from nothing to the correct conditions to run your column.

By putting them in a standardised sample-list that you use as a template, you won't have to go to the fiddle of adding them manually each time.