Hello to everyone out there!!! My name is Laura and I was wondering if anyone could help me. In order to scale up from analytical to prep for gradient time, the equation tp=tA X Fp/FA X Vp/VA
Where tA is run time of analytical, F = Flow, V= volume of column applies. Does this apply to isocratic runs as well. I am running an isocratic and gradient run together at the same time. How do I scale up for both at the same time???

Thanx

Consider the gradient portion and the isocratic portion of the run to be the exact same thing and scale both in exactly the same way. This measn that you need to scale the isocratic time according to the same formula as the gradient time. This will give you the same retention pattern in the prep run. The retention times may be slightly different due to the different influence of the gradient delay volume in the analytical and the prep run.

What is the best way to determine the dwell time (tD) for an HPLC system? Thanks!

Connect two containers with methanol to your solvent inlet lines. One container is spiked with somewhere around 0.5% acetone. Flush the lines, then run a step gradient (without column)from methanol to methanol with acetone. You'll get a step response in the UV. The 50% point is the gradient dwell time. You can also program this step after a preset time, and the difference between the preset time and the time at 50% in the detector gives you the same result.
There are other methods to do this as well, but I think this is the simplest method.

Hello to everyone again. I completed my purification runs using the scale up info for isocratic and gradient run. It worked well. Thank you very much Uwe.