Chromatography Forum

I'm trying to take a 3 mins isocratic run using conventional column and transfer it to UPLC to get a much shorter run time. However, when I use the UPLC calculator, the minimum retention factor I can enter is 5. Is there a way to enter less than 5 for the retention factor? What impact does retention factor has on the final conditions......other than N and run time? According to the UPLC, the retention factor is the analysis time divided by the column dead time minus 1. Can someone explain how you obtain this equation?

How curious, I just gave this same answer to another thread on this forum. We recently published an article in LCGC online that describes how to accelerate a method. The theory is rather tedious, so there is a step-by-step description and tutorial. See: http://www.nxtbook.com/nxtbooks/advanst ... tartpage=6

Hi gtma,

I never used the calculator so I don’t know whether there is a way of entering a smaller number than 5.
But I’m surprised by the fact that 5 is the smallest value one is allowed to enter. I always go for retention factor (K) between 2 and 10, when developing a new method. And I know many people who will agree that K ~ 2 is just fine. But as I said I find it easier to rely on gut feeling/experience. Especially in your case (isocratic elution) I wouldn’t bother to use the calculator.

K influences – besides N – the resolution and even the sensitivity. The latter is due to peak broadening, which results in lower peaks, when K is too high (e.g. > 10)

Her is the equation: K=(tR – t0)/t0 ~ (tR/t0) – 1

Best Regards

The scaling method is actually very straightforward, if you use a constant ratio of column length to particle size L/dp, for example going from a 15 cm 5 micron column to a 5 cm 1.7 micron column. Now, you simply need to increase the velocity by the ratio of particle sizes, in this case 5:1.7, and change the corresponding flow rate with the column crossection.

F(UPLC)/F(HPLC)= dp(HPLC)/dp(UPLC) * ID(UPLC)^2 / ID(HPLC)^2

dp is particle size, ID is column internal diameter.

The calculator will do this for you, even for a situation where the ratio of column length to particle size is not constant. It will do so by matching the plate count. The reason that I shied away from very short retention factors is that at short retention, extra-column effects play a role, and I can not calculate extra-column effects for an HPLC system that is unknown to me. The initial version of the calculator allowed the user to input extra-column effects, but based on user feedback, we threw this out again.

The calculation of the retention factor is in textbooks.

Mark, please excuse me, your post must have arrived while I wrote mine. I’m sure your link gives more comprehensive answers.

Best Regards